Anti-Inflammatory Compounds and Use Thereof

ABSTRACT

Compounds of the general formula 
     
       
         
         
             
             
         
       
     
     are disclosed with activity towards treating diseases related to inflammation, such as cancer, neurodegenerative and cardiovascular diseases. Pharmaceutical compositions and methods of use are also disclosed.

CROSS-REFERENCES TO RELATED APPLICATIONS

This application claims priority of U.S. Provisional Application Ser.No. 60/955,258, filed Aug. 10, 2007, and U.S. Provisional ApplicationSer. No. 60/989,584, filed Nov. 21, 2007. The contents of bothprovisional applications are incorporated herein by reference in theirentirety.

FIELD OF THE INVENTION

The invention is directed to compounds and pharmaceutical compositionsfor the treatment of inflammation-related diseases, in particularcancer.

BACKGROUND OF THE INVENTION

Inflammation, a key component of the immune system, functions in bothdefense and pathophysiological events to maintain the homeostasis oftissues, organs and individual cells. Inflammation can be classified aseither acute or chronic. Acute inflammation is a short-term processcharacterized by the classic signs of inflammation, i.e. swelling,redness, pain, heat, and loss of function, due to infiltration oftissues by plasma and leukocytes. It occurs as long as the injuriousstimulus is present and ceases once the stimulus has been removed.Chronic inflammation is a pathological condition characterized byconcurrent active inflammation, tissue destruction, and attempts atrepair. Chronically inflamed tissue is characterized by the infiltrationof mononuclear immune cells (monocytes, macrophages, lymphocytes, andplasma cells), tissue destruction, and attempts at healing, whichinclude angiogenesis and fibrosis.

Without inflammation, wounds and infections would not be able to healand progressive destruction of the tissue would threaten the survival ofthe organism. Unchecked inflammation, on the other hand, can lead to ahost of diseases, such as hay fever, atherosclerosis and othercardiovascular diseases, neurodegenerative diseases such as Alzheimer's,cancer and rheumatoid arthritis. For these reasons, inflammation istightly regulated by the body.

Inflammation is controlled by more than 400 genes. The pro-inflammatorygenotype, which appears dominant, increases our vulnerability to, andintensity of, inflammatory reactions, which underlie chronicinflammatory diseases, especially in old age. (Ferencik et al.,Inflammation—a lifelong companion. Folia Microbiol (Praha). 2007;52:159-73). Although joint diseases have long been the prototypicalinflammatory diseases, cardiovascular diseases, neurodegenerativediseases, autoimmune diseases, and cancer are now appreciated as havinginflammation as a unifying component of their pathogenesis.

Alzheimer's disease (AD) includes inflammatory processes in the senileplaques and surrounding glia, with increased expression of acute phaseproteins such as C-reactive protein (CRP) and IL-6. Increased IL-6expression during normal brain aging suggests a link of age-relatedinflammation to the onset of AD during aging. Blood levels of CRP andIL-6 are also associated with higher risk of Alzheimer's disease andcognitive decline during aging (Finch and Morgan, Systemic inflammation,infection, ApoE alleles, and Alzheimer disease: a position paper. CurrAlzheimer Res. 2007; 4:185-9).

Inflammation plays a crucial role in all steps characterizing theatherosclerotic process. Circulating CRP (C-reactive protein) levelshave emerged as a powerful independent determinant of cardiovascularevents. Hypertension is closely linked to inflammation. Experimentaldata and results from cross-sectional studies in humans strongly supportthis notion (Virdis et al., C-reactive protein and hypertension: isthere a causal relationship? Curr Pharm Des. 2007; 13:1693-8). Incancer, chronic inflammation often acts as a tumor promoter, resultingin aggressive cancerous growth and spread. Many of the same inflammatoryfactors that promote tumor growth also are responsible for cancercachexia/anorexia, pain, debilitation, and shortened survival. Acompelling case has been made even for attacking inflammation at initialdiagnosis to improving patient quality of life and survival. Serumlevels of CRP correlate with poor prognosis in cancer patients(MacDonald N. Cancer cachexia and targeting chronic inflammation: aunified approach to cancer treatment and palliative/supportive care. JSupport Oncol. 2007; 5:157-62).

Nonsteroidal anti-inflammatory drugs (NSAIDS) are the most widely usedanti-inflammatory compounds, with aspirin, the prototypical NSAID, stillbeing one of the oldest and most extensively used medication in theworld (Stanley P, Hegedus R. Aspirin—the first hundred years. Biologist(London) 2000; 47:269-71; Rinsema T J. One hundred years of aspirin. MedHist 1999; 43:502-7). NSAIDs have a significant antineoplastic effect,which should be viewed, at least in part, in the context of theincreasingly appreciated role of inflammation in cancer. Aspirin isformally documented to be a chemopreventive agent against colon cancer[3, 4]. For other NSAIDS, the evidence on their antineoplasticproperties is quite strong but still it is based mainly onepidemiological studies (Baron JA. What now for aspirin and cancerprevention? J Natl Cancer Inst 2004; 96:4-5; Jacobs E J, Rodriguez C,Mondul A M, Connell C J, Henley S J, Calle E E, et al. A large cohortstudy of aspirin and other nonsteroidal anti-inflammatory drugs andprostate cancer incidence. J Natl Cancer Inst 2005; 97:975-80; Thun M J,Henley S J, Gansler T. Inflammation and cancer: an epidemiologicalperspective. Novartis Foundation symposium 2004; 256:6-21; discussion2-8, 49-52, 266-9). For example, a recent meta-analysis of 91epidemiological studies showed a significant exponential decline withincreasing intake of NSAIDs in the risk for 7-10 malignancies includingthe four major types: colon, breast, lung, and prostate cancer (Harris RE, Beebe-Donk J, Doss H, Burr Doss D. Aspirin, ibuprofen, and othernon-steroidal anti-inflammatory drugs in cancer prevention: a criticalreview of non-selective COX-2 blockade (review). Oncology reports 2005;13:559-83; Ratliff TL. Aspirin, ibuprofen, and other non-steroidalanti-inflammatory drugs in cancer prevention: a critical review ofnon-selective COX-2 blockade (review). The Journal of urology 2005;174:787-8).

NSAIDs prevent cancer likely through pleiotropic effects (reviewed inRayyan Y, Williams J, Rigas B. The role of NSAIDs in the prevention ofcolon cancer. Cancer Invest 2002; 20:1002-11; Shiff S J, Rigas B.Aspirin for cancer. Nat Med 1999; 5:1348-9.). It is, however, clear thatconventional NSAIDs do not meet two important criteria for their wideapplication as chemopreventive agents against cancer, i.e. safety andhigh efficacy, as NSAIDs are associated with a considerable number ofside effects, and their efficacy is rather limited, not exceeding 50%(Rayyan Y, Williams J, Rigas B. The role of NSAIDs in the prevention ofcolon cancer. Cancer Invest 2002; 20:1002-11). Thus there is a need todevelop compounds with improved efficacy and safety profiles for thetreatment of various diseases related to inflammation.

SUMMARY OF THE INVENTION

The present invention provides such novel therapeutics including a novelgroup of NSAID derivatives and method of using them in the preventionand treatment of diseases, especially cancers.

In a first aspect, the present invention provides compounds of generalFormula I:

or an enantiomer, racemate, diastereomer, or tautomer thereof, or aprodrug, salt, hydrate or ester thereof;wherein X¹ is selected from the group consisting of —O— —S— and —NH—;wherein B is an optionally substituted aliphatic, alicyclic,heteroaliphatic, heterocyclic, aromatic, or heteroaromatic group,

wherein B is optionally substituted with one or more X² which isindependently selected from the group consisting of hydrogen, halogen,hydroxyl, alkoxyl or —CN; an optionally substituted aliphatic,alicyclic, heteroaliphatic, heterocyclic, aromatic, heteroaromaticmoiety; —OR^(R), —S(═O)_(n)R^(d), —NR^(b)R^(c), —C(═O)R^(a) and—C(═O)OR^(a); wherein n is 0-2, R^(R) is an optionally substitutedaliphatic, alicyclic, heteroaliphatic, heterocyclic, aryl or aralkyl,heteroaromatic group or acyl moiety;

R^(a), for each occurrence, is independently selected from the groupconsisting of hydrogen and an optionally substituted aliphatic,alicyclic, heteroaliphatic, heterocyclic, aromatic, or a heteroaromaticmoiety;

R^(b) and R^(c), for each occurrence, are independently selected fromthe group consisting of hydrogen; hydroxy; SO₂R^(d); and aliphatic,alicyclic, heteroaliphatic, heterocyclic, aromatic, heteroaromatic or anacyl moiety;

R^(d), for each occurrence, is independently selected from the groupconsisting of hydrogen; —N(R^(e))₂; aliphatic, aryl and heteroaryl; and

R^(e), for each occurrence, is independently hydrogen or an aliphaticgroup;

wherein A is an optionally substituted aliphatic, alicyclic,heteroaliphatic, heterocyclic, aromatic, or heteroaromatic group such asbut not limited to those moieties described in further detail hereinbelow;wherein D is hydroxyl; halide; tosylate; phosphate ester (—O—P(OR^(f))₃)or a phosphite ester (—O—P(OR^(g))₂), —OSO₂NR_(x)R_(y), where R_(x) andR_(y) are independently hydrogen, or a substituted or unsubstitutedaliphatic, alicyclic, heteroaliphatic, heterocyclic, aryl or aralkyl,heteroaromatic or acyl moiety; —O—C₆H₄OC(═O)CH₃; an alkoxy moiety; or anacyl moiety, provided that at least one R^(f) is not an H, and at leastone R^(g) is not an H, and provided that if B is

where R^(h) is aryl, aralkyl, alkyl, alkenyl or alkynyl, then D is aphosphate ester (—O—P(O)(OR^(f))₂) or a phosphite ester (—O—P(OR^(g))₂),preferably, R^(f) and R^(g) is independently each an H, alkyl, alkenyl,alkynyl, aryl or an aralkyl group, which may in turn be substituted orunsubstituted.

In one embodiment, the present invention relates to mono-, di- ortri-esters of phosphoric acid or phosphorous acid of aspirin, containingalkyl, alkenyl, aryl, benzyl or cyclic groups and their derivatives, andthe use thereof for cancer treatment and prevention. A series of novelaspirin derivatives, in particular 2-acetoxy-benzoic acid4-(diethoxy-phosphoryloxymethyl)-phenyl ester, and 2-acetoxy-benzoicacid 3-(diethoxy-phosphoryloxymethyl)-phenyl ester, provisionally namedphosphoaspirins, and determined these compounds have anticanceractivities both in vitro and in vivo. Phosphoaspirin inhibited thegrowth of HT-29 human colon adenocarcinoma cells (IC₅₀=276.6±12.3 μM(mean±SEM) through a combined antiproliferative and mainly proapoptoticeffect. While not willing to be bound by any theory on mechanism,applicant believes that phosphoaspirin achieves this effect bymodulating cell kinetics; the proliferation index of cancer cells wasreduced by 18.13% compared to controls (p<0.001) and the apoptosis indexwas increased by 94.6% (p<0.003). No apparent toxicity was shown byphosphoaspirin.

Preferably, compound of formula I is selected from the following:Diethyl 4-(2-acetoxylbenzoyloxy)benzyl phosphate, 2-acetoxy-benzoic acid4-(diethoxy-phosphoryloxymethyl)-phenyl ester, 2-acetoxy-benzoic acid3-(diethoxy-phosphoryloxymethyl)-phenyl ester, and phospho-sulindac I,phospho-sulindac II, phosphoflurbiprofen, phosphoibuprofen,glycero-phosphoaspirin I, and glycero-phosphoraspirin II. The chemicalstructures of these compounds are provided hereinbelow.

In a further aspect, the invention is directed to a pharmaceuticalcomposition comprising a compound of Formula I, as described generallyherein, and a pharmaceutically acceptable excipient. In a specificembodiment, the composition is useful in the treatment of human andanimal inflammation related diseases, including, but not limited toneoplasms, cancer. rheumatologic diseases such as rheumatoid arthritisand Sjogren's syndrome; cardiovascular diseases, such as coronary arterydisease, peripheral vascular disease and hypertension; neurodegenerativediseases, such as Alzheimer's disease and its variants orcerebrovascular diseases; and autoimmune diseases for example lupuserythematosus. Such compositions can comprise one or more otherpharmaceutical agents in addition to one or more compounds of theinvention.

In another embodiment, the invention is directed to a method forinhibiting inflammation, in particular chronic inflammation in a subjectin need thereof by administering to the subject an amount of thecompound or composition of the present invention effective to inhibitinflammation. The subject may be a human patient or animal.

In yet another aspect, the present invention provides methods fortreating any disorder related to undesirable inflammation comprisingadministering to a subject (e.g., human patient or animal) in needthereof a therapeutically effective amount of a compound of Formula I ofthe invention or a pharmaceutical composition comprising a compound ofthe invention. In a preferred embodiment, the disorder includes, but isnot limited to rheumatologic diseases such as for example rheumatoidarthritis and Sjogren's syndrome; cardiovascular diseases, such as, forexample, coronary artery disease, peripheral vascular disease andhypertension; neurodegenerative diseases, such as, for example,Alzheimer's disease and its variants or cerebrovascular diseases;autoimmune diseases such as for example lupus erythematosus; and otherconditions characterized by chronic inflammation of organs such as forexample the lung, such as chronic bronchitis or the sinuses, such aschronic sinusitis.

The compounds of the present invention may be used for the manufactureof a medicament for treatment of a disease listed above.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is the NMR profiles of several of the compounds of the presentinvention.

FIG. 2 shows that DFMO enhances phospho-sulindac-induced inhibition ofcolon cancer cell proliferation, as well as cell cycle arrest and celldeath by apoptosis. A—Left panel: Cell viability was determined in HT-29(full bars) and SW-480 (empty bars) cells after 48 h of incubation with5 mM DFMO, 40 μM phospho-sulindac (P-S) or simultaneous combination ofDFMO and P-S. Data are expressed as percentage of control cells (cellsincubated only with DMSO). Values are mean±SEM of 4 independentexperiments. *Significantly different from control cells (p<0.02, oneway ANOVA test). Right panel: In this isobologram the additivity lineconnects the IC₅₀ value of each compound used alone. A and B representtwo different dose pairs of each compound (their respectiveconcentrations are shown in parentheses). The location of both A and Bbelow the additivity line signifies synergy. B-Cell cycle progression inHT-29 cells incubated for 48 h with 5 mM DFMO, 40 μM phospho-sulindac(P-S) or simultaneous combination of DFMO and P-S. Representativeprofiles of the distribution of cells in G1, G2/M and S phases, out of 4independent experiments are shown. DNA content was determined frompropidium iodide (PI) fluorescence. C-Apoptosis and necrosis weredetermined by combined staining with annexin V and PI and determiningfluorescence intensity. The percentages of apoptotic cells weredetermined using the dual staining with annexin V and propidium iodideand are indicated in each quadrant: left bottom quadrant, viable cells(annexin V-negative/PI-negative); right bottom quadrant, early apoptoticcells (annexin V-positive/PI-negative); right upper quadrant, lateapoptotic cells (annexin V-positive/PI-positive); left upper quadrant,necrotic cells (annexin V-negative, PI-positive). These images arerepresentative of 3 independent experiments.

FIG. 3 shows that phospho-sulindac I inhibits NF-κB activation in coloncancer cells. Phospho-sulindac inhibits constitutive and TNFα-inducedNF-κB activation. Upper panel: Nuclear fractions were isolated fromHT-29 cells after 4 h of incubation in the absence or in the presence of40-100 μM phospho-sulindac I (P-S). Electrophoretic mobility shift assay(EMSA) for NF-κB and OCT-1 from cells treated with variousconcentrations of phospho-sulindac I. To determine the specificity ofeach transcription factor-DNA complex, the control nuclear fraction (−)was incubated in the presence of 100-fold molar excess of unlabeledoligonucleotide containing the consensus sequence for either thespecific (+S) or a nonspecific (+NS) transcription factor before thebinding assay. Lower panel: Nuclear fractions were isolated from HT-29cells, after 4 h of preincubation in the absence or in the presence of80 μM phospho-sulindac I (P-S) and further 0, 30 or 60 min incubationwithout (−) or with 10 ng/ml TNFα. EMSA for NF-κB incubated with variousconcentrations of phospho-sulindac I.

FIG. 4 shows that Phospho-sulindac I induces SSAT enzymatic activity incolon cancer cells. SW480 cells were incubated without or withphospho-sulindac I (P-S) for up to 24 h and SSAT activity was determinedat each time point by determining the amount of labelledN1-acetylspermidine synthesized from [¹⁴C]acetyl-CoA and unlabeledspermidine. Values are shown as means±SEM of 4 independent experiments.*Significantly different from control cells (p<0.01, one way ANOVAtest).

DETAILED DESCRIPTION OF THE INVENTION Definitions

The term “aliphatic”, as used herein, includes both saturated andunsaturated, straight chain (i.e., unbranched) or branched aliphatichydrocarbons, which are optionally substituted with one or morefunctional groups. As will be appreciated by one of ordinary skill inthe art, “aliphatic” is intended herein to include, but is not limitedto, alkyl, alkenyl, and alkynyl moieties. Thus, as used herein, the term“alkyl” includes straight and branched alkyl groups. An analogousconvention applies to other generic terms such as “alkenyl”, “alkynyl”and the like. Furthermore, as used herein, the terms “alkyl”, “alkenyl”,“alkynyl” and the like encompass both substituted and unsubstitutedgroups. In certain embodiments, as used herein, “lower alkyl” is used toindicate those alkyl groups (substituted, unsubstituted, branched orunbranched) having 1-6 carbon atoms.

In certain embodiments, the alkyl, alkenyl and alkynyl groups employedin the invention contain 1-20 aliphatic carbon atoms. In certain otherembodiments, the alkyl, alkenyl, and alkynyl groups employed in theinvention contain 1-10 aliphatic carbon atoms. In yet other embodiments,the alkyl, alkenyl, and alkynyl groups employed in the invention contain1-8 aliphatic carbon atoms. In still other embodiments, the alkyl,alkenyl, and alkynyl groups employed in the invention contain 1-6aliphatic carbon atoms. In yet other embodiments, the alkyl, alkenyl,and alkynyl groups employed in the invention contain 1-4 carbon atoms.Illustrative aliphatic groups thus include, but are not limited to, forexample, methyl, ethyl, n-propyl, isopropyl, allyl, n-butyl, sec-butyl,isobutyl, tert-butyl, n-pentyl, sec-pentyl, isopentyl, tert-pentyl,n-hexyl, sec-hexyl, moieties and the like, which again, may bear one ormore substituents. Alkenyl groups include, but are not limited to, forexample, ethenyl, propenyl, butenyl, 1-methyl-2-buten-1-yl, and thelike. Representative alkynyl groups include, but are not limited to,ethynyl, 2-propynyl (propargyl), 1-propynyl and the like.

The term “alicyclic”, as used herein, refers to compounds, which combinethe properties of aliphatic and cyclic compounds and include but are notlimited to monocyclic, or polycyclic aliphatic hydrocarbons and bridgedcycloalkyl compounds, which are optionally substituted with one or morefunctional groups. As will be appreciated by one of ordinary skill inthe art, “alicyclic” is intended herein to include, but is not limitedto, cycloalkyl, cycloalkenyl, and cycloalkynyl moieties, which areoptionally substituted with one or more functional groups. Illustrativealicyclic groups thus include, but are not limited to, for example,cyclopropyl, —CH₂-cyclopropyl, cyclobutyl, —CH₂-cyclobutyl, cyclopentyl,—CH₂-cyclopentyl, cyclohexyl, —CH₂-cyclohexyl, cyclohexenylethyl,cyclohexanylethyl, norborbyl moieties and the like, which again, maybear one or more substituents.

The term “alkoxy” or “alkyloxy”, as used herein refers to a saturated(i.e., O-alkyl) or unsaturated (i.e., O-alkenyl and O-alkynyl) groupattached to the parent molecular moiety through an oxygen atom. Incertain embodiments, the alkyl group contains 1-20 aliphatic carbonatoms. In certain other embodiments, the alkyl group contains 1-10aliphatic carbon atoms. In yet other embodiments, the alkyl, alkenyl,and alkynyl groups employed in the invention contain 1-8 aliphaticcarbon atoms. In still other embodiments, the alkyl group contains 1-6aliphatic carbon atoms. In yet other embodiments, the alkyl groupcontains 1-4 aliphatic carbon atoms. Examples of alkoxy, include but arenot limited to, methoxy, ethoxy, propoxy, isopropoxy, n-butoxy,i-butoxy, sec-butoxy, tert-butoxy, neopentoxy, n-hexoxy and the like.

The term “alkylthio” or “thioalkyl” as used herein refers to a saturated(i.e., S-alkyl) or unsaturated (i.e., S-alkenyl and S-alkynyl) groupattached to the parent molecular moiety through a sulfur atom. Incertain embodiments, the alkyl group contains 1-20 aliphatic carbonatoms. In certain other embodiments, the alkyl group contains 1-10aliphatic carbon atoms. In yet other embodiments, the alkyl, alkenyl,and alkynyl groups employed in the invention contain 1-8 aliphaticcarbon atoms. In still other embodiments, the alkyl group contains 1-6aliphatic carbon atoms. In yet other embodiments, the alkyl groupcontains 1-4 aliphatic carbon atoms. Examples of thioalkyl include, butare not limited to, methylthio, ethylthio, propylthio, isopropylthio,n-butylthio, and the like.

The term “alkylamino” refers to a group having the structure—NHR′wherein R′ is alkyl, as defined herein. The term “aminoalkyl” refers toa group having the structure NH₂R′—, wherein R′ is alkyl, as definedherein. In certain embodiments, the alkyl group contains 1-20 aliphaticcarbon atoms. In certain other embodiments, the alkyl group contains1-10 aliphatic carbon atoms. In yet other embodiments, the alkyl,alkenyl, and alkynyl groups employed in the invention contain 1-8aliphatic carbon atoms. In still other embodiments, the alkyl groupcontains 1-6 aliphatic carbon atoms. In yet other embodiments, the alkylgroup contains 1-4 aliphatic carbon atoms. Examples of alkylaminoinclude, but are not limited to, methylamino, ethylamino,iso-propylamino and the like.

Some examples of substituents of the above-described aliphatic (andother) moieties of compounds of the invention include, but are notlimited to aliphatic; alicyclic; heteroaliphatic; heterocyclic;aromatic; heteroaromatic; aryl; heteroaryl; alkylaryl; heteroalkylaryl;alkylheteroaryl; heteroalkylheteroaryl; alkoxy; aryloxy; heteroalkoxy;heteroaryloxy; alkylthio; arylthio; heteroalkylthio; heteroarylthio; F;C1; Br; I; —OH; —NO₂; —ONO₂; —CN; —CF₃; —CH₂CF₃; —CHC1₂; —CH₂OH;—CH₂CH₂OH; —CH₂NH₂; —CH₂SO₂CH₃; —C(O)R_(x); —CO₂(R_(x)); —CON(R_(x))₂;—OC(O)R_(x); —OCO₂R_(x); —OCON(R_(x))₂; —N(R_(x))₂; —S(O)₂R_(x);—NR_(x)(CO)R_(x) wherein each occurrence of R_(x) independentlyincludes, but is not limited to, aliphatic, alicyclic, heteroaliphatic,heterocyclic, aryl, heteroaryl, alkylaryl, alkylheteroaryl,heteroalkylaryl or heteroalkylheteroaryl, wherein any of the aliphatic,alicyclic, heteroaliphatic, heterocyclic, alkylaryl, or alkylheteroarylsubstituents described above and herein may be substituted orunsubstituted, branched or unbranched, saturated or unsaturated, andwherein any of the aryl or heteroaryl substituents described above andherein may be substituted or unsubstituted. Additional examples ofgenerally applicable substituents are illustrated by the specificembodiments shown in the Examples that are described herein.

In general, the term “aromatic moiety”, as used herein, refers to astable mono- or polycyclic, unsaturated moiety having preferably 3-14carbon atoms, each of which may be substituted or unsubstituted. Incertain embodiments, the term “aromatic moiety” refers to a planar ringhaving p-orbitals perpendicular to the plane of the ring at each ringatom and satisfying the Huckel rule where the number of pi electrons inthe ring is (4n+2) wherein n is an integer. A mono- or polycyclic,unsaturated moiety that does not satisfy one or all of these criteriafor aromaticity is defined herein as “non-aromatic”, and is encompassedby the term “alicyclic.”

In general, the term “heteroaromatic moiety”, as used herein, refers toa stable mono- or polycyclic, unsaturated moiety having preferably 3-14carbon atoms, each of which may be substituted or unsubstituted; andcomprising at least one heteroatom selected from O, S and N within thering (i.e., in place of a ring carbon atom). In certain embodiments, theterm “heteroaromatic moiety” refers to a planar ring comprising at leastone heteroatom, having p-orbitals perpendicular to the plane of the ringat each ring atom, and satisfying the Huckel rule where the number of pielectrons in the ring is (4n+2) wherein n is an integer.

It will also be appreciated that aromatic and heteroaromatic moieties,as defined herein may be attached via an alkyl or heteroalkyl moiety andthus also include -(alkyl)aromatic, -(heteroalkyl)aromatic,-(heteroalkyl)heteroaromatic, and -(heteroalkyl)heteroaromatic moieties.Thus, as used herein, the phrases “aromatic or heteroaromatic moieties”and “aromatic, heteroaromatic-(alkyl)aromatic, -(heteroalkyl)aromatic,-(heteroalkyl)heteroaromatic, and -(heteroalkyl)heteroaromatic” areinterchangeable. Substituents include, but are not limited to, any ofthe previously mentioned substituents, i.e., the substituents recitedfor aliphatic moieties, or for other moieties as disclosed herein,resulting in the formation of a stable compound.

The term “aryl”, as used herein, does not differ significantly from thecommon meaning of the term in the art, and refers to an unsaturatedcyclic moiety comprising at least one aromatic ring. In certainembodiments, “aryl” refers to a mono- or bicyclic carbocyclic ringsystem having one or two aromatic rings including, but not limited to,phenyl, naphthyl, tetrahydronaphthyl, indanyl, indenyl and the like.

The term “heteroaryl”, as used herein, does not differ significantlyfrom the common meaning of the term in the art, and refers to a cyclicaromatic radical having from five to ten ring atoms of which one ringatom is selected from S, O and N; zero, one or two ring atoms areadditional heteroatoms independently selected from S, O and N; and theremaining ring atoms are carbon, the radical being joined to the rest ofthe molecule via any of the ring atoms, such as, for example, pyridyl,pyrazinyl, pyrimidinyl, pyrrolyl, pyrazolyl, imidazolyl, thiazolyl,oxazolyl, isooxazolyl, thiadiazolyl, oxadiazolyl, thiophenyl, furanyl,quinolinyl, isoquinolinyl, and the like.

It will be appreciated that aryl and heteroaryl groups (includingbicyclic aryl groups) can be unsubstituted or substituted, whereinsubstitution includes replacement of one or more of the hydrogen atomsthereon independently with any one or more of the following moietiesincluding, but not limited to: aliphatic; alicyclic; heteroaliphatic;heterocyclic; aromatic; heteroaromatic; aryl; heteroaryl; alkylaryl;heteroalkylaryl; alkylheteroaryl; heteroalkylheteroaryl; alkoxy;aryloxy; heteroalkoxy; heteroaryloxy; alkylthio; arylthio;heteroalkylthio; heteroarylthio; F; C1; Br; I; —OH; —NO₂; —CN; —CF₃;—CH₂CF₃; —CHC1₂; —CH₂OH; —CH₂CH₂OH; —CH₂NH₂; —CH₂SO₂CH₃; —C(O)R_(x);—CO₂(R_(x)); —CON(R_(X))₂; —OC(O)R_(x); —OCO₂R_(x); —OCON(R_(x))₂;—N(R_(x))₂; —S(O)R_(x); —S(O)₂R_(x); —NR_(x)(CO)R_(x) wherein eachoccurrence of R_(x) independently includes, but is not limited to,aliphatic, alicyclic, heteroaliphatic, heterocyclic, aromatic,heteroaromatic, aryl, heteroaryl, alkylaryl, alkylheteroaryl,heteroalkylaryl or heteroalkylheteroaryl, wherein any of the aliphatic,alicyclic, heteroaliphatic, heterocyclic, alkylaryl, or alkylheteroarylsubstituents described above and herein may be substituted orunsubstituted, branched or unbranched, saturated or unsaturated, andwherein any of the aromatic, heteroaromatic, aryl, heteroaryl,-(alkyl)aryl or -(alkyl)heteroaryl substituents described above andherein may be substituted or unsubstituted. Additionally, it will beappreciated, that any two adjacent groups taken together may represent a4, 5, 6, or 7-membered substituted or unsubstituted alicyclic orheterocyclic moiety. Additional examples of generally applicablesubstituents are illustrated by the specific embodiments shown in theExamples that are described herein.

The term “cycloalkyl”, as used herein, refers specifically to groupshaving three to seven, preferably three to ten carbon atoms. Suitablecycloalkyls include, but are not limited to cyclopropyl, cyclobutyl,cyclopentyl, cyclohexyl, cycloheptyl and the like, which, as in the caseof aliphatic, alicyclic, heteroaliphatic or heterocyclic moieties, mayoptionally be substituted with substituents including, but not limitedto aliphatic; alicyclic; heteroaliphatic; heterocyclic; aromatic;heteroaromatic; aryl; heteroaryl; alkylaryl; heteroalkylaryl;alkylheteroaryl; heteroalkylheteroaryl; alkoxy; aryloxy; heteroalkoxy;heteroaryloxy; alkylthio; arylthio; heteroalkylthio; heteroarylthio; F;C1; Br; I; —OH; —NO₂; —CN; —CF₃; —CH₂CF₃; —CHC1₂; —CH₂OH; —CH₂CH₂OH;—CH₂NH₂; —CH₂SO₂CH₃; —C(O)R_(x); —CO₂(R_(x)); —CON(R_(x))₂; —OC(O)R_(x);—OCO₂R_(x); —OCON(R_(x))₂; —N(R_(x))₂; —S(O)₂R_(x); —NR_(x)(CO)R_(x)wherein each occurrence of R_(x) independently includes, but is notlimited to, aliphatic, alicyclic, heteroaliphatic, heterocyclic,aromatic, heteroaromatic, aryl, heteroaryl, alkylaryl, alkylheteroaryl,heteroalkylaryl or heteroalkylheteroaryl, wherein any of the aliphatic,alicyclic, heteroaliphatic, heterocyclic, alkylaryl, or alkylheteroarylsubstituents described above and herein may be substituted orunsubstituted, branched or unbranched, saturated or unsaturated, andwherein any of the aromatic, heteroaromatic, aryl or heteroarylsubstituents described above and herein may be substituted orunsubstituted. Additional examples of generally applicable substituentsare illustrated by the specific embodiments shown in the Examples thatare described herein.

The term “heteroaliphatic”, as used herein, refers to aliphatic moietiesin which one or more carbon atoms in the main chain have beensubstituted with a heteroatom. Thus, a heteroaliphatic group refers toan aliphatic chain which contains one or more oxygen, sulfur, nitrogen,phosphorus or silicon atoms, e.g., in place of carbon atoms.Heteroaliphatic moieties may be linear or branched, and saturated orunsaturated. In certain embodiments, heteroaliphatic moieties aresubstituted by independent replacement of one or more of the hydrogenatoms thereon with one or more moieties including, but not limited toaliphatic; alicyclic; heteroaliphatic; heterocyclic; aromatic;heteroaromatic; aryl; heteroaryl; alkylaryl; alkylheteroaryl; alkoxy;aryloxy; heteroalkoxy; heteroaryloxy; alkylthio; arylthio;heteroalkylthio; heteroarylthio; F; C1; Br; I; —OH; —NO₂; —CN; —CF₃;—CH₂CF₃; —CHC1₂; —CH₂OH; —CH₂CH₂OH; —CH₂NH₂; —CH₂SO₂CH₃; —C(O)R_(x);—CO₂(R_(x)); —CON(R_(x))₂; —OC(O)R_(x); —OCO₂R_(x); —OCON(R_(x))₂;—N(R_(x))₂; —S(O)₂R_(x); —NR_(x)(CO)R_(x) wherein each occurrence ofR_(x) independently includes, but is not limited to, aliphatic,alicyclic, heteroaliphatic, heterocyclic, aromatic, heteroaromatic,aryl, heteroaryl, alkylaryl, alkylheteroaryl, heteroalkylaryl orheteroalkylheteroaryl, wherein any of the aliphatic, alicyclic,heteroaliphatic, heterocyclic, alkylaryl, or alkylheteroarylsubstituents described above and herein may be substituted orunsubstituted, branched or unbranched, saturated or unsaturated, andwherein any of the aromatic, heteroaromatic, aryl or heteroarylsubstituents described above and herein may be substituted orunsubstituted. Additional examples of generally applicable substituentsare illustrated by the specific embodiments shown in the Examples thatare described herein.

The term “heterocycloalkyl”, “heterocycle” or “heterocyclic”, as usedherein, refers to compounds which combine the properties ofheteroaliphatic and cyclic compounds and include, but are not limitedto, saturated and unsaturated mono- or polycyclic cyclic ring systemshaving 5-16 atoms wherein at least one ring atom is a heteroatomselected from O, S and N (wherein the nitrogen and sulfur heteroatomsmay be optionally be oxidized), wherein the ring systems are optionallysubstituted with one or more functional groups, as defined herein. Incertain embodiments, the term “heterocycloalkyl”, “heterocycle” or“heterocyclic” refers to a non-aromatic 5-, 6- or 7-membered ring or apolycyclic group wherein at least one ring atom is a heteroatom selectedfrom O, S and N (wherein the nitrogen and sulfur heteroatoms may beoptionally be oxidized), including, but not limited to, a bi- ortri-cyclic group, comprising fused six-membered rings having between oneand three heteroatoms independently selected from oxygen, sulfur andnitrogen, wherein (i) each 5-membered ring has 0 to 2 double bonds, each6-membered ring has 0 to 2 double bonds and each 7-membered ring has 0to 3 double bonds, (ii) the nitrogen and sulfur heteroatoms may beoptionally be oxidized, (iii) the nitrogen heteroatom may optionally bequaternized, and (iv) any of the above heterocyclic rings may be fusedto an aryl or heteroaryl ring. Representative heterocycles include, butare not limited to, heterocycles such as furanyl, thiofuranyl, pyranyl,pyrrolyl, thienyl, pyrrolidinyl, pyrazolinyl, pyrazolidinyl,imidazolinyl, imidazolidinyl, piperidinyl, piperazinyl, oxazolyl,oxazolidinyl, isooxazolyl, isoxazolidinyl, dioxazolyl, thiadiazolyl,oxadiazolyl, tetrazolyl, triazolyl, thiatriazolyl, oxatriazolyl,thiadiazolyl, oxadiazolyl, morpholinyl, thiazolyl, thiazolidinyl,isothiazolyl, isothiazolidinyl, dithiazolyl, dithiazolidinyl,tetrahydrofuryl, and benzofused derivatives thereof. In certainembodiments, a “substituted heterocycle, or heterocycloalkyl orheterocyclic” group is utilized and as used herein, refers to aheterocycle, or heterocycloalkyl or heterocyclic group, as definedabove, substituted by the independent replacement of one, two or threeof the hydrogen atoms thereon with but are not limited to aliphatic;alicyclic; heteroaliphatic; heterocyclic; aromatic; heteroaromatic;aryl; heteroaryl; alkylaryl; heteroalkylaryl; alkylheteroaryl;heteroalkylheteroaryl; alkoxy; aryloxy; heteroalkoxy; heteroaryloxy;alkylthio; arylthio; heteroalkylthio; heteroarylthio; F; Cl; Br; I; —OH;—NO₂; —CN; —CF₃; —CH₂CF₃; —CHCl₂; —CH₂OH; —CH₂CH₂OH; —CH₂NH₂;—CH₂SO₂CH₃; —C(O)R_(x); —CO₂(R_(x)); —CON(R_(x))₂; —OC(O)R_(x);—OCO₂R_(x); —OCON(R_(x))₂; —N(R_(x))₂; —S(O)₂R_(x); —NR_(x)(CO)R_(x)wherein each occurrence of R_(x) independently includes, but is notlimited to, aliphatic, alicyclic, heteroaliphatic, heterocyclic,aromatic, heteroaromatic, aryl, heteroaryl, alkylaryl, alkylheteroaryl,heteroalkylaryl or heteroalkylheteroaryl, wherein any of the aliphatic,alicyclic, heteroaliphatic, heterocyclic, alkylaryl, or alkylheteroarylsubstituents described above and herein may be substituted orunsubstituted, branched or unbranched, saturated or unsaturated, andwherein any of the aromatic, heteroaromatic, aryl or heteroarylsubstituents described above and herein may be substituted orunsubstituted. Additional examples or generally applicable substituentsare illustrated by the specific embodiments shown in the Examples, whichare described herein.

Additionally, it will be appreciated that any of the alicyclic orheterocyclic moieties described above and herein may comprise an aryl orheteroaryl moiety fused thereto. Additional examples of generallyapplicable substituents are illustrated by the specific embodimentsshown in the Examples that are described herein.

The terms “halo” “halide” and “halogen” as used herein refer to an atomselected from fluorine, chlorine, bromine and iodine.

The term “haloalkyl” denotes an alkyl group, as defined above, havingone, two, or three halogen atoms attached thereto and is exemplified bysuch groups as chloromethyl, bromoethyl, trifluoromethyl, and the like.

The term “amino”, as used herein, refers to a primary (—NH₂), secondary(—NHR_(x)), tertiary (—NR_(x)R_(y)) or quaternary (—N⁺R_(x)R_(y)R_(z))amine, where R_(x), R_(y) and R_(z) are independently an aliphatic,alicyclic, heteroaliphatic, heterocyclic, aromatic or heteroaromaticmoiety, as defined herein. Examples of amino groups include, but are notlimited to, methylamino, dimethylamino, ethylamino, diethylamino,diethylaminocarbonyl, methylethylamino, iso-propylamino, piperidino,trimethylamino, and propylamino.

The term “acyl”, as used herein, refers to a group having the generalformula —C(═O)R, where R is an aliphatic, alicyclic, heteroaliphatic,heterocyclic, aromatic or heteroaromatic moiety, as defined herein.

The term “sulfonamido”, as used herein, refers to a group of the generalformula —SO₂NR_(x)R_(y), where R_(x) and R_(y) are independentlyhydrogen, or an aliphatic, alicyclic, heteroaliphatic, heterocyclic,aromatic, heteroaromatic or acyl moiety, as defined herein.

The term “benzamido”, as used herein, refers to a group of the generalformula PhCONR_(x)—, where R_(x) is hydrogen, or an aliphatic,alicyclic, heteroaliphatic, heterocyclic, aromatic, heteroaromatic oracyl moiety, as defined herein.

The term “C₁₋₆alkylidene”, as used herein, refers to a substituted orunsubstituted, linear or branched saturated divalent radical consistingsolely of carbon and hydrogen atoms, having from one to six carbonatoms, having a free valence “—” at both ends of the chain.

The term “C₂₋₆alkenylidene”, as used herein, refers to a substituted orunsubstituted, linear or branched unsaturated divalent radicalconsisting solely of carbon and hydrogen atoms, having from two to sixcarbon atoms, having a free valence “—” at both ends of the radical, andwherein the unsaturation is present only as double bonds and wherein adouble bond can exist between the first carbon of the chain and the restof the molecule.

As used herein, the terms “aliphatic”, “heteroaliphatic”, “alkyl”,“alkenyl”, “alkynyl”, “heteroalkyl”, “heteroalkenyl”, “heteroalkynyl”,and the like encompass substituted and unsubstituted, saturated andunsaturated, and linear and branched groups. Similarly, the terms“alicyclic”, “heterocyclic”, “heterocycloalkyl”, “heterocycle” and thelike encompass substituted and unsubstituted, and saturated andunsaturated groups. Additionally, the terms “cycloalkyl”,“cycloalkenyl”, “cycloalkynyl”, “heterocycloalkyl”,“heterocycloalkenyl”, “heterocycloalkynyl”, “aromatic”,“heteroaromatic”, “aryl”, “heteroaryl” and the like encompass bothsubstituted and unsubstituted groups.

The phrase, “pharmaceutically acceptable derivative”, as used herein,denotes any pharmaceutically acceptable salt, ester, or salt of suchester, of such compound, or any other adduct or derivative which, uponadministration to a patient, is capable of providing (directly orindirectly) a compound as otherwise described herein, or a metabolite orresidue thereof. Pharmaceutically acceptable derivatives thus includeamong others pro-drugs. A pro-drug is a derivative of a compound,usually with significantly reduced pharmacological activity, whichcontains at least one additional moiety, which is susceptible to removalin vivo yielding the parent molecule as the pharmacologically activespecies. An example of a pro-drug is an ester, which is cleaved in vivoto yield a compound of interest. Pro-drugs of a variety of compounds,and materials and methods for derivatizing the parent compounds tocreate the pro-drugs, are known and may be adapted to the presentinvention. Certain exemplary pharmaceutical compositions andpharmaceutically acceptable derivatives will be discussed in more detailherein below.

The present invention discloses compounds and pharmaceuticalcompositions thereof that possess anti-inflammatory activities.

The compounds of the invention include compounds of the general formula(I) as defined below:

or a stereoisomer such as an enantiomer or a diastereomer or a racemate,or a tautomer thereof, or a prodrug, salt, hydrate or ester thereof;wherein X′ is selected from the group consisting of —O—, —S—, and —NH—;wherein B is an optionally substituted aliphatic, alicyclic,heteroaliphatic, heterocyclic, aryl, aralkyl or a heteroaromatic group,and wherein B is optionally substituted by one or more substituents X²which is independently selected from the group consisting of hydrogen,halogen, hydroxyl, —NO₂, —ONO₂, —CN; an optionally substitutedaliphatic, alicyclic, heteroaliphatic, heterocyclic, aromatic,heteroaromatic moiety; —OR^(R), —S(═O)_(n)R^(d), —NR^(b)R^(c),—C(═O)R^(a) and —C(═O)OR^(a); wherein n is 0-2, R^(R) is an optionallysubstituted aliphatic, alicyclic, heteroaliphatic, heterocyclic,aromatic, heteroaromatic or acyl moiety;R^(a), for each occurrence, is independently selected from the groupconsisting of hydrogen and an optionally substituted aliphatic,alicyclic, heteroaliphatic, heterocyclic, aromatic, or a heteroaromaticmoiety;R^(b) and R^(c), for each occurrence, are independently selected fromthe group consisting of hydrogen; hydroxy; SO₂R^(d); and aliphatic,alicyclic, heteroaliphatic, heterocyclic, aromatic, heteroaromatic or anacyl moiety;R^(d), for each occurrence, is independently selected from the groupconsisting of hydrogen; —N(R^(e))₂; aliphatic, aryl and heteroaryl; andR^(e), for each occurrence, is independently hydrogen or aliphatic;wherein A is an optionally substituted aliphatic, alicyclic,heteroaliphatic, heterocyclic, aromatic, or heteroaromatic group such asbut not limited to those moieties described in further detail hereinbelow;wherein D is hydroxyl; halide; tosylate; phosphate ester(—O—P(O)(OR^(f))₂) or a phosphite ester (—O—P(OR^(g))₂),—OSO₂NR_(x)R_(y), where R_(x) and R_(y) are independently hydrogen, or asubstituted or unsubstituted aliphatic, alicyclic, heteroaliphatic,heterocyclic, aromatic, heteroaromatic or acyl moiety; —O—C₆H₄OC(═O)CH₃;an alkoxy moiety; or an acyl moiety, provided that at least one R^(f) isnot an H, and at least one R^(g) is not an H, and provided that if B is

where R^(h) is aryl, aralkyl, alkyl, alkenyl or alkynyl, then D isphosphate ester (—O—P(OR^(f))₃) or a phosphite ester (—O—P(OR^(g))₂),preferably, R^(f) and R^(g) is independently each an H, alkyl, alkenylor an alkynyl group, which may in turn be substituted or unsubstituted.

The compounds of Formula I are not limited by the position of thesubstituents on an aromatic ring. For example, if B is an aromatic ring,the -D moiety may be meta, ortho or para to the A-C(═O)—X¹ moiety, inparticular when X² is H. If one or more X² substituents are present,they may be positioned at any unoccupied position(s). Thus, any and allpositional isomers of compounds of Formula I are embraced by theinvention. As will be apparent from the further discussion below onsynthetic methods for the compounds of the invention, the A-C(═O)—X¹—moiety is facilely derived from a carboxylic acid-containing reactant(A-C(═O)—OH) or an amide-containing reactant (A-C(═O)—NH), and thus theA-C(═O)—X¹— moiety may be referred to herein as being derived from acompound with the structure A-C(═O)—OH or A-C(═O)—NH.

In one embodiment of compounds of Formula (I) of the invention, A is

wherein X² is one or more substituents as defined above, and Y is(—C—)_(n), wherein n is 0 to 4, and when n is 2 or more, Y optionallycontains one or more unsaturated bonds. For example, when n=0, theoptionally substituted aromatic ring is bonded to the —C(═O)—X¹—substituent of Formula (I). When n=1, Y is —CH₂—. When n=2, Y may be—CH₂—CH₂—, —CH═CH— or ethynyl radical. When n=3, Y may be —CH₂—CH₂—CH₂—,an allyl radical, —CH═CH—CH₂— or —CH₂—CH═CH—, or a triple bond withinthe radical. When n=4, the divalent radical may have any combination ofsaturation and unsaturation.

Among the preferred but non-limiting selections of substituent A ofFormula I, in a first embodiment, A is derived from among non-steroidalanti-inflammatory drugs (NSAIDs) including but not limited to aspirin,sulindac, ibuprofen, flurbiprofen, or formula IV or an analog of eitherof the foregoing.

Suitable analogs of formula IV include but are not limited toderivatives with one or more fluorine atoms substituted on one or bothof the benzene rings of the formula IV moiety; and compounds with one ormore substitutions on the alpha carbon, such as ethyl, dimethyl,diethyl, propyl and other such aliphatic substitutions. Thus, in oneembodiment, A may be

wherein X² is one or more substituents as described above, R² is atleast one halogen, and R₃ and R₄ are independently hydrogen or analiphatic group. In a preferred embodiment, R² is F. In a more preferredembodiment, X² is H, R² is F (at position 3 relative to CR³R⁴) and R³and R⁴ are H and CH₃, respectively.Thus, in one preferred but non-limiting embodiment, A can be

In a second embodiment, A is derived from aspirin, such as shown below:

where X² is one or more substituents as described above.

In a preferred but non-limiting embodiment, A is

In a third embodiment, A is derived from cinnamic acid, or an analog ofcinnamic acid, such as is shown below:

where R⁵ and R⁶ are independently hydrogen, —OH, alkoxy, halide,trifluoroalkyl, alpha-haloalkyl, trifluoroalkoxy, or R^(a) as describedabove. Non-limiting examples of the foregoing include trifluoromethyl,alpha-fluoromethyl, 4-(anisylideneamino), 2-(hexadecyloxy), and4-nitro-alpha-(ortho-tolyl). Examples of Formula VI from which group Ain Formula I can be selected include but are not limited to3,4-dihydroxy, o-, m- and p-hydroxy; 2,3-dihydroxy; 3,5-dihydroxy;3,4-dimethoxy; 3-hydroxy-4-methoxy and 3,4-dimethoxy. Thus, A can be

In another embodiment, A is derived from phthalic acid, or an analog ofphthalic acid, shown below:

wherein R⁵ and R⁶ are as described above. Examples of such A moietiesinclude:

In yet a further embodiment, A is a straight chain or branched aliphaticmoiety, preferably 1 to 7 carbons. In compounds wherein A is analiphatic group, X² is preferably a moiety derived from theesterification of resveratrol or an analog thereof to a carboxylic acidon the aromatic ring, i.e. X² is (—C═O)OR^(a). Suitable analogs ofresveratrol include but are not limited to the compounds described byShe Q-B et al. in Oncogene, volume 22, pp 2143-2150, 2003, and in thepublication by Roberti et al. in J. Med Chem, volume 46, pp 3546-3554,2003. In one embodiment, X² is

wherein R⁵ and R⁶ are as described above.

Non-limiting selections of X² are thus, by way of non-limiting examples,

In a preferred embodiment of the foregoing, A is methyl.

In addition to the foregoing selections of A, also embraced by theinvention are compounds of Formula I wherein A is selected from thegroup consisting of an optionally substituted aliphatic, alicyclic,heteroaliphatic, aromatic, heterocyclic or heteroaromatic moiety.

The D substituent of Formula I is hydroxyl; halide; tosylate; phosphateester (—O—P(O)(OR^(f))₂) or a phosphite ester (—O—P(OR^(g))₂),—OSO₂NR_(x)R_(y), where R_(x) and R_(y) are independently hydrogen, or asubstituted or unsubstituted aliphatic, alicyclic, heteroaliphatic,heterocyclic, aromatic, heteroaromatic or acyl moiety; —O—C₆H₄OC(═O)CH₃;an alkoxy moiety; or an acyl moiety, provided that at least one R^(f) isnot an H, and at least one R^(g) is not an H.

In the compounds of the present invention according to formula I, if Bis

where R^(h) is aryl, aralkyl, alkyl, alkenyl or alkynyl then D isphosphate ester (—O—P(O)(OR^(f))₂) or a phosphite ester (—O—P(OR^(g))₂),wherein R^(f) and R^(g) is independently each an H, alkyl, alkenyl or analkynyl group, which may in turn be substituted or unsubstituted. The Dsubstituent of Formula I is preferably a phosphate ester or a phosphiteester moiety, such as —OPO(alkyloxy)₂, —OPO₂(alkyloxy), —OP(alkyloxy)₂,—OPO(alkyloxy).

As noted above, in case B contains a benzene ring, the substituentcontaining the aforementioned D moiety, —CH²⁻-D, may be at any locationon the benzene ring relative to the position of the —X₁—C(═O)-Asubstituent, i.e., meta, ortho or para thereto. The invention embracesall such positional isomers.

The selections among substituent X² are as described above. As mentionedabove, in certain cases where A is an aliphatic group such as methyl, X²may be a carboxylic acid to which an alcohol or polyphenol isesterified, such as resveratrol or an analog thereof. Suitable analogsof resveratrol include but are not limited to the compounds described byShe Q-B et al. in Oncogene, volume 22, pp 2143-2150, 2003, and in thepublication by Roberti et al. in J. Med. Chem., volume 46, pp 3546-3554,2003. Other preferred examples of X² include one or more —OH, —OCH₃, or—F, at one or more positions not occupied by the substituents containingmoieties A and D. Other preferred examples of X² include —CH₃, and—C₂H₅.

Preferably, compound of formula I is selected from the following:2-acetoxy-benzoic acid 4-(diethoxy-phosphoryloxymethyl)-phenyl ester,2-acetoxy-benzoic acid 3-(diethoxy-phosphoryloxymethyl)-phenyl ester,and phospho-sulindac I, phospho-sulindac II, phosphoflurbiprofen,phosphoibuprofen, phosphoaspirin I, phosphoraspirin II, andphospho-valproic acid, as described hereinbelow.

The foregoing compounds are merely illustrative of Formula I and are notintended to be limiting.

In a further aspect, the invention is directed to a pharmaceuticalcomposition comprising a compound of Formula I, as described generallyherein, and a pharmaceutically acceptable excipient. In a specificembodiment, the composition is useful in the treatment of human andanimal inflammation related diseases including but not limited toneoplasms and cancer, rheumatologic diseases such as rheumatoidarthritis and Sjogren's syndrome; cardiovascular diseases, such ascoronary artery disease, peripheral vascular disease and hypertension;neurodegenerative diseases such as Alzheimer's disease and its variantsor cerebrovascular diseases; and autoimmune diseases such as lupuserythematosus; and other conditions characterized by chronicinflammation of organs such as the lung, such as chronic bronchitis orthe sinuses, such as chronic sinusitis; cardiovascular diseases, forexample, coronary artery disease, peripheral vascular disease andhypertension; neurodegenerative diseases, for example, Alzheimer'sdisease and its variants or cerebrovascular diseases; and autoimmunediseases such as lupus erythematosus; other conditions characterized bychronic inflammation of organs such as the lung, such as chronicbronchitis or the sinuses, such as chronic sinusitis; and variousneoplastic and pre-neoplastic diseases, for example, benign prostatichypertrophy, prostate cancer, colon adenomas and colon cancer, cancer ofthe lung, lymphomas and leukemias.

Such compositions can comprise one or more other pharmaceutical agentsin addition to one or more compounds of the invention.

In another embodiment, the invention is directed to a method forinhibiting inflammation, in particular chronic inflammation in a subjectin need thereof by administering to the subject an amount of thecompound or composition of the present invention effective to inhibitinflammation. The subject may be a human patient or animal.

It will be appreciated that for each of the classes and subclassesdescribed above and herein, any one or more occurrences of aliphatic orheteroaliphatic may independently be substituted or unsubstituted,cyclic or acyclic, linear or branched and any one or more occurrences ofaryl, heteroaryl, cycloaliphatic, cycloheteroaliphatic may besubstituted or unsubstituted.

Some of the foregoing compounds can comprise one or more asymmetriccenters, and thus can exist in various isomeric forms, e.g.,stereoisomers and/or diastereomers. Thus, inventive compounds andpharmaceutical compositions thereof may be in the form of an individualenantiomer, diastereomer or geometric isomer, or may be in the form of amixture of stereoisomers. In certain embodiments, the compounds of theinvention are enantiopure compounds. In certain other embodiments,mixtures of stereoisomers or diastereomers are provided. Moreover, whencompounds of the invention exist in tautomeric forms, each tautomer isembraced herein.

Furthermore, certain compounds, as described herein may have one or moredouble bonds that can exist as either the Z or E isomer, unlessotherwise indicated. The invention additionally encompasses thecompounds as individual isomers substantially free of other isomers andalternatively, as mixtures of various isomers, e.g., racemic mixtures ofstereoisomers. In addition to the above-mentioned compounds per se, thisinvention also encompasses pharmaceutically acceptable derivatives ofthese compounds and compositions comprising one or more compounds of theinvention and one or more pharmaceutically acceptable excipients oradditives.

Thus, the invention is directed to the use of the aforementionedcompounds for treating inflammation-related diseases.

Thus, in a specific embodiment, the invention is directed to a methodfor obtaining a pharmaceutical composition, comprising formulating thecompounds of the present invention into a composition comprising thecompound of the present invention and a pharmaceutically acceptablecarrier or excipient. The invention is further directed to uses of thecompound of the present invention for manufacturing a medicament.

Compositions

As discussed above, this invention provides novel compounds that havebiological properties useful for the treatment of any of a number ofconditions or diseases generally characterized by abnormal inflammation,or prophylaxis in instances wherein a risk of appearance of suchconditions or diseases is present. Moreover, certain compounds known inthe art have been newly identified as having activity likewise useful inthe prophylaxis or treatment of abnormal inflammation, and the inventionis also directed to anti-inflammation compositions comprising suchcompounds.

Accordingly, in another aspect of the present invention, pharmaceuticalcompositions are provided, which comprise any one of the compoundsdescribed herein (or a prodrug, pharmaceutically acceptable salt orother pharmaceutically acceptable derivative thereof), and optionallycomprise a pharmaceutically acceptable carrier. In certain embodiments,these compositions optionally further comprise one or more additionaltherapeutic agents. Alternatively, a compound of this invention may beadministered to a patient in need thereof in combination with theadministration of one or more other therapeutic agents. For example,additional therapeutic agents for conjoint administration or inclusionin a pharmaceutical composition with a compound of this invention may bean approved anti-inflammation agent, or it may be any one of a number ofagents undergoing approval in the Food and Drug Administration thatultimately obtain approval for the treatment of any disorder related toinflammation. Such additional therapeutic agents may also be provided topromote the targeting of the compounds of the invention to the desiredsite of treatment, or may increase its stability, increase itshalf-life, etc. It will also be appreciated that certain of thecompounds of present invention can exist in free form for treatment, orwhere appropriate, as a pharmaceutically acceptable derivative thereof.According to the present invention, a pharmaceutically acceptablederivative includes, but is not limited to, pharmaceutically acceptablesalts, esters, salts of such esters, or a pro-drug or other adduct orderivative of a compound of this invention which upon administration toa patient in need is capable of providing, directly or indirectly, acompound as otherwise described herein, or a metabolite or residuethereof.

As used herein, the term “pharmaceutically acceptable salt” refers tothose salts which are, within the scope of sound medical judgment,suitable for use in contact with the tissues of humans and lower animalswithout undue toxicity, irritation, allergic response and the like, andare commensurate with a reasonable benefit/risk ratio. Pharmaceuticallyacceptable salts of amines, carboxylic acids, and other types ofcompounds, are well known in the art. For example, S. M. Berge, et al.describe pharmaceutically acceptable salts in detail in J.Pharmaceutical Sciences, 66: 1-19 (1977), incorporated herein byreference. The salts can be prepared in situ during the final isolationand purification of the compounds of the invention, or separately byreacting a free base or free acid function with a suitable reagent, asdescribed generally below. For example, a free base function can bereacted with a suitable acid. Furthermore, where the compounds of theinvention carry an acidic moiety, suitable pharmaceutically acceptablesalts thereof may, include metal salts such as alkali metal salts, e.g.sodium or potassium salts; and alkaline earth metal salts, e.g. calciumor magnesium salts. Examples of pharmaceutically acceptable, nontoxicacid addition salts are salts of an amino group formed with inorganicacids such as hydrochloric acid, hydrobromic acid, phosphoric acid,sulfuric acid and perchloric acid or with organic acids such as aceticacid, oxalic acid, maleic acid, tartaric acid, citric acid, succinicacid or malonic acid or by using other methods used in the art such asion exchange. Other pharmaceutically acceptable salts include adipate,alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate,borate, butyrate, camphorate, camphorsulfonate, citrate,cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate,formate, fumarate, glucoheptonate, glycerophosphate, gluconate,hemisulfate, heptanoate, hexanoate, hydroiodide,2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, laurylsulfate, malate, maleate, malonate, methanesulfonate,2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate,pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate,pivalate, propionate, stearate, succinate, sulfate, tartrate,thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and thelike. Representative alkali or alkaline earth metal salts includesodium, lithium, potassium, calcium, magnesium, and the like. Furtherpharmaceutically acceptable salts include, when appropriate, nontoxicammonium, quaternary ammonium, and amine cations formed usingcounterions such as halide, hydroxide, carboxylate, sulfate, phosphate,nitrate, loweralkyl sulfonate and aryl sulfonate.

Additionally, as used herein, the term “pharmaceutically acceptableester” refers to esters that hydrolyze in vivo and include those thatbreak down readily in the human body to leave the parent compound or asalt thereof. Suitable ester groups include, for example, those derivedfrom pharmaceutically acceptable aliphatic carboxylic acids,particularly alkanoic, alkenoic, cycloalkanoic and alkanedioic acids, inwhich each alkyl or alkenyl moiety advantageously has not more than 6carbon atoms. Examples of particular esters include formates, acetates,propionates, butyrates, acrylates and ethylsuccinates.

Furthermore, the term “pharmaceutically acceptable prodrugs” as usedherein refers to those prodrugs of the compounds of the presentinvention which are, within the scope of sound medical judgment,suitable for use in contact with the issues of humans and lower animalswith undue toxicity, irritation, allergic response, and the like,commensurate with a reasonable benefit/risk ratio, and effective fortheir intended use, as well as the zwitterionic forms, where possible,of the compounds of the invention. The term “prodrug” refers tocompounds that are rapidly transformed in vivo to yield the parentcompound of the above formula, for example by hydrolysis in blood. Athorough discussion is provided in T. Higuchi and V. Stella, Pro-drugsas Novel Delivery Systems, Vol. 14 of the A.C.S. Symposium Series, andin Edward B. Roche, ed., Bioreversible Carriers in Drug Design, AmericanPharmaceutical Association and Pergamon Press, 1987, both of which areincorporated herein by reference.

As described above, the pharmaceutical compositions of the presentinvention additionally comprise a pharmaceutically acceptable carrier,which, as used herein, includes any and all solvents, diluents, or otherliquid vehicle, dispersion or suspension aids, surface active agents,isotonic agents, thickening or emulsifying agents, preservatives, solidbinders, lubricants and the like, as suited to the particular dosageform desired. Remington's Pharmaceutical Sciences, Sixteenth Edition, E.W. Martin (Mack Publishing Co., Easton, Pa., 1980) discloses variouscarriers used in formulating pharmaceutical compositions and knowntechniques for the preparation thereof. Except insofar as anyconventional carrier medium is incompatible with the compounds of theinvention, such as by producing any undesirable biological effect orotherwise interacting in a deleterious manner with any othercomponent(s) of the pharmaceutical composition, its use is contemplatedto be within the scope of this invention. Some examples of materialswhich can serve as pharmaceutically acceptable carriers include, but arenot limited to, sugars such as lactose, glucose and sucrose; starchessuch as corn starch and potato starch; cellulose and its derivativessuch as sodium carboxymethyl cellulose, ethyl cellulose and celluloseacetate; powdered tragacanth; malt; gelatine; talc; excipients such ascocoa butter and suppository waxes; oils such as peanut oil, cottonseedoil; safflower oil, sesame oil; olive oil; corn oil and soybean oil;glycols; such as propylene glycol; esters such as ethyl oleate and ethyllaurate; agar; buffering agents such as magnesium hydroxide and aluminumhydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer'ssolution; ethyl alcohol, and phosphate buffer solutions, as well asother non-toxic compatible lubricants such as sodium lauryl sulfate andmagnesium stearate, as well as coloring agents, releasing agents,coating agents, sweetening, flavoring and perfuming agents,preservatives and antioxidants can also be present in the composition,according to the judgment of the formulator.

Liquid dosage forms for oral administration include, but are not limitedto, pharmaceutically acceptable emulsions, microemulsions, solutions,suspensions, syrups and elixirs. In addition to the active compounds,the liquid dosage forms may contain inert diluents commonly used in theart such as, for example, water or other solvents, solubilizing agentsand emulsifiers such as ethyl alcohol, isopropyl alcohol, ethylcarbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propyleneglycol, 1,3-butylene glycol, dimethylformamide, oils (in particular,cottonseed, groundnut (peanut), corn, germ, olive, castor, and sesameoils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols andfatty acid esters of sorbitan, and mixtures thereof. Besides inertdiluents, the oral compositions can also include adjuvants such aswetting agents, emulsifying and suspending agents, sweetening,flavoring, and perfuming agents.

Injectable preparations, for example, sterile injectable aqueous oroleaginous suspensions may be formulated according to the known artusing suitable dispersing or wetting agents and suspending agents. Thesterile injectable preparation may also be a sterile injectablesolution, suspension or emulsion in a nontoxic parenterally acceptablediluent or solvent, for example, as a solution in 1,3-butanediol. Amongthe acceptable vehicles and solvents that may be employed are water,Ringer's solution, U.S.P. and isotonic sodium chloride solution. Inaddition, sterile, fixed oils are conventionally employed as a solventor suspending medium. For this purpose, any bland fixed oil can beemployed including synthetic mono- or diglycerides. In addition, fattyacids such as oleic acid are used in the preparation of injectables.

The injectable formulations can be sterilized, for example, byfiltration through a bacterial-retaining filter, or by incorporatingsterilizing agents in the form of sterile solid compositions which canbe dissolved or dispersed in sterile water or other sterile injectablemedium prior to use.

In order to prolong the effect of a drug, it is often desirable to slowthe absorption of the drug from subcutaneous or intramuscular injection.This may be accomplished by the use of a liquid suspension orcrystalline or amorphous material with poor water solubility. The rateof absorption of the drug then depends upon its rate of dissolutionthat, in turn, may depend upon crystal size and crystalline form.Alternatively, delayed absorption of a parenterally administered drugform is accomplished by dissolving or suspending the drug in an oilvehicle. Injectable depot forms are made by forming microencapsulematrices of the drug in biodegradable polymers such aspolylactide-polyglycolide. Depending upon the ratio of drug to polymerand the nature of the particular polymer employed, the rate of drugrelease can be controlled. Examples of other biodegradable polymersinclude poly(orthoesters) and poly(anhydrides). Depot injectableformulations are also prepared by entrapping the drug in liposomes ormicroemulsions which are compatible with body tissues.

Compositions to deliver the agent directly to the colon—for example,pills from which the active agent is released into the colon by apH-dependent or other mechanism ensuring exclusive or predominantcolonic delivery of said compound, suppositories, enemas and other meansfor colonic delivery.

Compositions for rectal or vaginal administration are preferablysuppositories which can be prepared by mixing the compounds of thisinvention with suitable non-irritating excipients or carriers such ascocoa butter, polyethylene glycol or a suppository wax which are solidat ambient temperature but liquid at body temperature and therefore meltin the rectum or vaginal cavity and release the active compound.

Solid dosage forms for oral administration include but are not limitedto capsules, tablets, pills, powders, and granules. In such solid dosageforms, the active compound is mixed with at least one inert,pharmaceutically acceptable excipient or carrier such as sodium citrateor dicalcium phosphate and/or a) fillers or extenders such as starches,lactose, sucrose, glucose, mannitol, and silicic acid, b) binders suchas, for example, carboxymethylcellulose, alginates, gelatin,polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such asglycerol, d) disintegrating agents such as agar-agar, calcium carbonate,potato or tapioca starch, alginic acid, certain silicates, and sodiumcarbonate, e) solution retarding agents such as paraffin, f) absorptionaccelerators such as quaternary ammonium compounds, g) wetting agentssuch as, for example, cetyl alcohol and glycerol monostearate, h)absorbents such as kaolin and bentonite clay, and i) lubricants such astalc, calcium stearate, magnesium stearate, solid polyethylene glycols,sodium lauryl sulfate, and mixtures thereof. In the case of capsules,tablets and pills, the dosage form may also comprise buffering agents.

Solid compositions of a similar type may also be employed as fillers insoft and hard-filled gelatin capsules using such excipients as lactoseor milk sugar as well as high molecular weight polyethylene glycols andthe like. The solid dosage forms of tablets, dragees, capsules, pills,and granules can be prepared with coatings and shells such as entericcoatings and other coatings well known in the pharmaceutical formulatingart. They may optionally contain opacifying agents and can also be of acomposition that they release the active ingredient(s) only, orpreferentially, in a certain part of the intestinal tract, optionally,in a delayed manner. Examples of embedding compositions that can be usedinclude polymeric substances and waxes. Solid compositions of a similartype may also be employed as fillers in soft and hard-filled gelatincapsules using such excipients as lactose or milk sugar as well as highmolecular weight polyethylene glycols and the like.

The active compounds can also be in micro-encapsulated form with one ormore excipients as noted above. The solid dosage forms of tablets,dragees, capsules, pills, and granules can be prepared with coatings andshells such as enteric coatings, release controlling coatings and othercoatings well known in the pharmaceutical formulating art. In such soliddosage forms, the active compound may be admixed with at least one inertdiluent such as sucrose, lactose and starch. Such dosage forms may alsocomprise, as in normal practice, additional substances other than inertdiluents, e.g., tableting lubricants and other tableting aids such asmagnesium stearate and microcrystalline cellulose. In the case ofcapsules, tablets and pills, the dosage forms may also comprisebuffering agents. They may optionally contain opacifying agents and canalso be of a composition that they release the active ingredient(s)only, or preferentially, in a certain part of the intestinal tract,optionally, in a delayed manner. Examples of embedding compositionswhich can be used include but are not limited to polymeric substancesand waxes.

The present invention encompasses pharmaceutically acceptable topicalformulations of inventive compounds. The term “pharmaceuticallyacceptable topical formulation”, as used herein, means any formulationwhich is pharmaceutically acceptable for intradermal administration of acompound of the invention by application of the formulation to theepidermis. In certain embodiments of the invention, the topicalformulation comprises a carrier system. Pharmaceutically effectivecarriers include, but are not limited to, solvents (e.g., alcohols, polyalcohols, water), creams, lotions, ointments, oils, plasters, liposomes,powders, emulsions, microemulsions, and buffered solutions (e.g.,hypotonic or buffered saline) or any other carrier known in the art fortopically administering pharmaceuticals. A more complete listing ofart-known carriers is provided by reference texts that are standard inthe art, for example, Remington's Pharmaceutical Sciences, 16^(th)Edition, 1980 and 17^(th) Edition, 1985, both published by MackPublishing Company, Easton, Pa., the disclosures of which areincorporated herein by reference in their entireties. In certain otherembodiments, the topical formulations of the invention may compriseexcipients. Any pharmaceutically acceptable excipient known in the artmay be used to prepare the inventive pharmaceutically acceptable topicalformulations. Examples of excipients that can be included in the topicalformulations of the invention include, but are not limited to,preservatives, antioxidants, moisturizers, emollients, buffering agents,solubilizing agents, other penetration agents, skin protectants,surfactants, and propellants, and/or additional therapeutic agents usedin combination to the inventive compound. Suitable preservativesinclude, but are not limited to, alcohols, quaternary amines, organicacids, parabens, and phenols. Suitable antioxidants include, but are notlimited to, ascorbic acid and its esters, sodium bisulfite, butylatedhydroxytoluene, butylated hydroxyanisole, tocopherols, and chelatingagents like EDTA and citric acid. Suitable moisturizers include, but arenot limited to, glycerin, sorbitol, polyethylene glycols, urea, andpropylene glycol. Suitable buffering agents for use with the inventioninclude, but are not limited to, citric, hydrochloric, and lactic acidbuffers. Suitable solubilizing agents include, but are not limited to,quaternary ammonium chlorides, cyclodextrins, benzyl benzoate, lecithin,and polysorbates. Suitable skin protectants that can be used in thetopical formulations of the invention include, but are not limited to,vitamin E oil, allatoin, dimethicone, glycerin, petrolatum, and zincoxide.

In certain embodiments, the pharmaceutically acceptable topicalformulations of the invention comprise at least a compound of theinvention and a penetration enhancing agent. The choice of topicalformulation will depend or several factors, including the condition tobe treated, the physicochemical characteristics of the inventivecompound and other excipients present, their stability in theformulation, available manufacturing equipment, and costs constraints.As used herein the term “penetration enhancing agent” means an agentcapable of transporting a pharmacologically active compound through thestratum corneum and into the epidermis or dermis, preferably, withlittle or no systemic absorption. A wide variety of compounds have beenevaluated as to their effectiveness in enhancing the rate of penetrationof drugs through the skin. See, for example, Percutaneous PenetrationEnhancers, Maibach H. I. and Smith H. E. (eds.), CRC Press, Inc., BocaRaton, Fla. (1995), which surveys the use and testing of various skinpenetration enhancers, and Buyuktimkin et al., Chemical Means ofTransdermal Drug Permeation Enhancement in Transdermal and Topical DrugDelivery Systems, Gosh T. K., Pfister W. R., Yum S. I. (Eds.),Interpharm Press Inc., Buffalo Grove, III. (1997). In certain exemplaryembodiments, penetration agents for use with the invention include, butare not limited to, triglycerides (e.g., soybean oil), aloe compositions(e.g., aloe-vera gel), ethyl alcohol, isopropyl alcohol,octolyphenylpolyethylene glycol, oleic acid, polyethylene glycol 400,propylene glycol, N-decylmethylsulfoxide, fatty acid esters (e.g.,isopropyl myristate, methyl laurate, glycerol monooleate, and propyleneglycol monooleate) and N-methyl pyrrolidone.

In certain embodiments, the compositions may be in the form ofointments, pastes, creams, lotions, gels, powders, solutions, sprays,inhalants or patches. In certain exemplary embodiments, formulations ofthe compositions according to the invention are creams, which mayfurther contain saturated or unsaturated fatty acids such as stearicacid, palmitic acid, oleic acid, palmito-oleic acid, cetyl or oleylalcohols, stearic acid being particularly preferred. Creams of theinvention may also contain a non-ionic surfactant, for example,polyoxy-40-stearate. In certain embodiments, the active component isadmixed under sterile conditions with a pharmaceutically acceptablecarrier and any needed preservatives or buffers as may be required.Ophthalmic formulation, eardrops, and eye drops are also contemplated asbeing within the scope of this invention. Additionally, the presentinvention contemplates the use of transdermal patches, which have theadded advantage of providing controlled delivery of a compound to thebody. Such dosage forms are made by dissolving or dispensing thecompound in the proper medium. As discussed above, penetration enhancingagents can also be used to increase the flux of the compound across theskin. The rate can be controlled by either providing a rate controllingmembrane or by dispersing the compound in a polymer matrix or gel.

It will also be appreciated that the compounds and pharmaceuticalcompositions of the present invention can be formulated and employed incombination therapies, that is, the compounds and pharmaceuticalcompositions can be formulated with or administered concurrently with,prior to, or subsequent to, one or more other desired therapeutics ormedical procedures. The particular combination of therapies(therapeutics or procedures) to employ in a combination regimen willtake into account compatibility of the desired therapeutics and/orprocedures and the desired therapeutic effect to be achieved. It willalso be appreciated that the therapies employed may achieve a desiredeffect for the same disorder (for example, an inventive compound may beadministered concurrently with another anti-inflammation agent), or theymay achieve different effects (e.g., control of any adverse effects).

In certain embodiments, the pharmaceutical compositions of the presentinvention further comprise one or more additional therapeutically activeingredients (e.g., anti-inflammatory and/or palliative). For purposes ofthe invention, the term “Palliative” refers to treatment that is focusedon the relief of symptoms of a disease and/or side effects of atherapeutic regimen, but is not curative. For example, palliativetreatment encompasses painkillers, antinausea medications andanti-sickness drugs.

In certain embodiments the compounds of the present invention can becovalently or non-covalently bound to for example polyethylene glycol orother similar molecules to make them suitable for administration to thepatient either in one of the forms described above or using nanodevices.Alternatively, the compounds of the present invention can be formulatedusing the principles of nanoscience to optimize their therapeuticapplication.

Uses and Methods of Treatment

As discussed above, certain of the compounds as described herein exhibitactivity generally as inhibitors of inflammation, with inflammationunderstood as described herein under “BACKGROUND OF THE INVENTION.”Thus, in certain embodiments, compounds of the invention are useful forthe treatment of any of a number of conditions or diseases in whichinflammation, in particular chronic inflammation is the cause of orrelates to the onset or continued occurrence of the disease orcondition, such as but not limited to rheumatologic diseases such asrheumatoid arthritis and Sjogren's syndrome; cardiovascular diseases,for example, coronary artery disease, peripheral vascular disease andhypertension; neurodegenerative diseases, for example, Alzheimer'sdisease and its variants or cerebrovascular diseases; and autoimmunediseases such as lupus erythematosus; other conditions characterized bychronic inflammation of organs such as the lung, such as chronicbronchitis or the sinuses, such as chronic sinusitis.

Accordingly, in another aspect of the invention, methods for thetreatment of inflammation-related disorders are provided comprisingadministering a therapeutically effective amount of a compound ofFormula I to a subject in need thereof. In certain embodiments, a methodfor the treatment of related disorders is provided comprisingadministering a therapeutically effective amount of an inventivecompound, or a pharmaceutical composition comprising an inventivecompound to a subject in need thereof, in such amounts and for such timeas is necessary to achieve the desired result.

The invention is also directed to the use of any compound of Formula (I)for the preparation of a medicament for administration to a human oranimal patient in need thereof, to inhibit or block inflammation. Suchcompounds preferably are administered once an inflammation-relateddisease or an inflammatory condition that may predispose to disease hasbeen diagnosed in the patient, optionally in combination with otheranti-inflammation agents or other agents such as those that maintaintherapeutic levels of the compounds within the body. Compounds of theinvention also may be administered after other therapies have been triedand failed, and may be administered prophylactically.

In certain embodiments, the uses and methods of the invention involvethe administration of a therapeutically effective amount of the compoundor a pharmaceutically acceptable derivative thereof to a subject(including, but not limited to a human or animal, including livestock,domesticated or zoo animals) in need thereof.

It will be appreciated that the compounds and compositions, according tothe method of the present invention, may be administered using anyamount and any route of administration effective for the treatment ofconditions or diseases in which anti-inflammation or related activitieshave a therapeutically useful role. Thus, the expression “effectiveamount” as used herein, refers to a sufficient amount of agent toinhibit inflammation and to exhibit a therapeutic effect. The exactamount required will vary from subject to subject, depending on thespecies, age, and general condition of the subject, the severity of theinfection, the particular therapeutic agent, its mode of administration,and the like. The compounds of the invention are preferably formulatedin dosage unit form for ease of administration and uniformity of dosage.The expression “dosage unit form” as used herein refers to a physicallydiscrete unit of therapeutic agent appropriate for the patient to betreated. It will be understood, however, that the total daily usage ofthe compounds and compositions of the present invention will be decidedby the attending physician within the scope of sound medical judgment.The specific therapeutically effective dose level for any particularpatient or organism will depend upon a variety of factors including thedisorder being treated and the severity of the disorder; the activity ofthe specific compound employed; the specific composition employed; theage, body weight, general health, sex and diet of the patient; the timeof administration, route of administration, and rate of excretion of thespecific compound employed; the duration of the treatment; drugs used incombination or coincidental with the specific compound employed; andlike factors well known in the medical arts.

Furthermore, after formulation with an appropriate pharmaceuticallyacceptable carrier in a desired dosage, the pharmaceutical compositionsof this invention can be administered to humans and other animalsorally, rectally, parenterally, intracisternally, intravaginally,intraperitoneally, topically (as by powders, ointments, or drops),bucally, as an oral or nasal spray, or the like, depending on thelocation and extent of the disease being treated. In certainembodiments, the compounds of the invention may be parenterallyadministered at dosage levels of about 0.001 mg/kg to about 50 mg/kg,from about 0.01 mg/kg to about 25 mg/kg, or from about 0.1 mg/kg toabout 10 mg/kg of subject body weight per day, one or more times a day,to obtain the desired therapeutic effect. In other embodiments,compounds of the invention may be administered orally or rectally atdosage levels of about 0.01 mg/kg to about 100 mg/kg, from about 0.05mg/kg to about 50 mg/kg, or from about 0.1 mg/kg to about 10 mg/kg ofsubject body weight per day, one or more times a day, to obtain thedesired therapeutic effect. It will also be appreciated that dosagessmaller than 0.001 mg/kg or greater than 50 mg/kg (for example 50-100mg/kg) can be administered to a subject. In certain embodiments,compounds are administered orally or parenterally.

Treatment Kit

In other embodiments, the present invention relates to a kit forconveniently and effectively carrying out the methods in accordance withthe present invention. In general, the pharmaceutical pack or kitcomprises one or more containers filled with one or more of theingredients of the pharmaceutical compositions of the invention. Suchkits are especially suited for the delivery of solid oral forms such astablets or capsules. Such a kit preferably includes a number of unitdosages, and may also include a card having the dosages oriented in theorder of their intended use. If desired, a memory aid can be provided,for example in the form of numbers, letters, or other markings or with acalendar insert, designating the days in the treatment schedule in whichthe dosages can be administered. Alternatively, placebo dosages, orcalcium dietary supplements, either in a form similar to or distinctfrom the dosages of the pharmaceutical compositions, can be included toprovide a kit in which a dosage is taken every day. Optionallyassociated with such container(s) can be a notice in the form prescribedby a governmental agency regulating the manufacture, use or sale ofpharmaceutical products, which notice reflects approval by the agency ofmanufacture, use or sale for human administration.

The representative examples that follow are intended to help illustratethe invention, and are not intended to, nor should they be construed to,limit the scope of the invention. Indeed, various modifications of theinvention and many further embodiments thereof, in addition to thoseshown and described herein, will become apparent to those skilled in theart from the full contents of this document, including the exampleswhich follow and the references to the scientific and patent literaturecited herein. It should further be appreciated that the contents ofthose cited references are incorporated herein by reference to helpillustrate the state of the art.

The following examples contain important additional information,exemplification and guidance that can be adapted to the practice of thisinvention in its various embodiments and the equivalents thereof.

EXAMPLES Example 1 Method of Synthesis

The following reaction scheme was followed to obtain Compound 5(para-phosphoaspirin) of this invention shown below. Compound 5 wassynthesized starting from O-Acetylsalicyloyl chloride (1) and4-hydroxybenzaldehyde (2) in three steps, as shown below.

In this scheme —OEt represents CH₃CH₂O—.

Step 1: Preparation of Compound 3

To a pre-cooled (O° C.) solution of 4-hydroxybenzaldehyde (2, 1.04 g,8.49 mmole) in dichloromethane (10 mL) and pyridine (4.16 mL, 50 mmole)was added O-acetylsalicyloyl chloride (1, 1.98 g, 10 mmole) in methylenechloride (10 mL) drop-wise between 0-5° C. The temperature of thereaction mixture was slowly raised to room temperature and left overnight. At this point TLC of the reaction mixture showed the completionof the reaction. The reaction mixture was washed with water (25 mL),followed by 1N HCl (25 mL) and then finally with aqueous NaHCO₃. Theorganic layer was dried over anhydrous sodium sulfate, filtered andconcentrated. The crude weight of the oil was 2.35 g (97%).

Step 2: Preparation of Compound 4

To a pre-cooled (O° C.) solution of compound 3 (2.3 g, 8.1 mmole) inmethylene chloride (10 mL) and acetic acid (2.5 mL) was added sodiumcyanoborohydride (253 mg, 4 mmole) in two portions. The temperature ofthe reaction was slowly raised to room temperature in 30 minutes. Atthis point TLC showed the completion of the reaction. The reactionmixture was washed with water (2×25 mL), followed by saturated aqueoussodium bicarbonate (25 mL) and then finally with brine. It was driedover anhydrous sodium sulfate, filtered and concentrated. The crudeweight of the solid was 1.95 g (83%).

Step 3: Preparation of 2-Acetoxy-benzoic acid4-(diethoxy-phosphoryloxymethyl)-phenyl ester (5)

To a solution of alcohol (4, 1.9 g, 6.64 mmole) in methylene chloride(10 mL) and diisopropylethylamine (2.2 mL, 13.28 mmole) was addeddiethylchlorophosphate (2.5 mL, 17.26 mmole) drop-wise, followed by DMAP(25 mg) as a solid. The reaction mixture was heated under refluxovernight. At this point the TLC showed the completion of the reaction.The reaction mixture was washed with water (2×25 mL), dried overanhydrous sodium sulfate, filtered and concentrated. The crude residuewas purified by column chromatography using hexane:ethyl acetate(60:40). The pure fractions were combined and evaporated to give a solidwhich was triturated with hot hexane several times to give pure titlecompound 690 mg (25%) as a solid.

To confirm purity and identity of compound 5 of this invention, TLC and¹H NMR was performed. The NMR profile is shown in FIG. 1.

NSAID Based Compounds

Non-steroidal anti-inflammatory drugs (NSAIDs) comprise a structurallyand, to a large extent, functionally diverse group of compounds withnearly 50 individual compounds approved for the treatment of patientswith a variety of inflammatory diseases. They all have analgesic,antipyretic and anti-inflammatory effects. Some of them, such asacetylsalicylic acid (aspirin), have been demonstrated to have an effectagainst inflammation-related diseases such as rheumatologic,cardiovascular, neurodegenerative and cancer, be this effect eithertherapeutic as, for example, in rheumatoid arthritis, or preventive as,for example, in cancer, coronary artery disease or Alzheimer's disease.

Broadly, NSAIDs can be categorized into the following chemical groups:salicylates, arylalkanoic acids (e.g. sulindac), 2-arylpropionic acids(profens), N-arylanthranilic acids (fenamic acids), pyrazolidinederivatives, oxicams, and sulphonanilides. Most of the available NSAIDsare amenable to derivatization as described herein using methods readilyavailable and known to those ordinarily skilled in the art.

The following are examples of several derivatized NSAIDs according tocertain embodiments of the invention.

The following compounds have been similarly synthesized.

Note: —OEt represents CH₃CH₂O-.asdfad

Two derivatives of sulindac(2-[6-fluoro-2-methyl-3-[(4-methylsulfinylphenyl)methylidene]inden-1-yl]-acetic acid) were synthesized. They are referredto as phospho-sulindac I and phsphosulindac II, respectively. Theirstructures are:

and their respective ¹H-NMR profiles are shown in FIG. 1.

One derivative of ibuprofen has been synthesized and is referred to asphospho-ibuprofen. Its structure is:

and its ¹H-NMR profile is shown in FIG. 1.

One derivative of flurbiprofen has been synthesized and is referred toas phospho-flurbiprofen. Its structure is:

and its ¹H-NMR profile is shown in FIG. 1.

Two derivatives of aspirin were synthesized. They are referred to asglycero-phospho-aspirin I and glycero-phospho-aspirin II, respectively.Their structures are shown below, and their respective ¹H-NMR profilesare shown in FIG. 1.

A derivative of naproxen is shown below

Example 4 Six Novel NSAID-Based Compounds have Significantly EnhancedAntineoplastic Potency Compared to their Parent Compounds

Six compounds based on four representative NSAIDs, aspirin (2derivatives differing in their linker), ibuprofen, flurbiprofen andsulindac (2 derivatives, differing in the structure of the sulindacmoiety) were synthesized following the methodology of Penning et al(Penning T D, Talley J J, Bertenshaw S R, Carter J S, Collins P W,Docter S, et al. Synthesis and biological evaluation of the1,5-diarylpyrazole class of cyclooxygenase-2 inhibitors: identificationof4-[5-(4-methylphenyl)-3-(trifluoromethyl)-1H-pyrazol-1-yl]benzenesulfonamide(SC-58635, celecoxib). J Med Chem 1997; 40:1347-65.) and the methodsdescribed herein. Their structures and NMR profiles are shown above.

Conventional NSAIDs were purchased from Sigma (St Louis, Mo.). Weexamined these six compounds for their antineoplastic properties,determined in cultured human cells derived from colon, breast andpancreatic cancers.

Cell Culture:

Human breast (MCF-7 and MDA-MB 231), colon (HT-29, and SW-480) andpancreatic (MIA PaCa-2 and BxPC-3) cell lines (American Type CultureCollection, Manassas, Va.) were grown as monolayers in the specificmedium suggested by American Type Culture Collection and supplementedwith 10% fetal calf serum (Mediatech, Herndon, Va.), penicillin (50U/ml) and streptomycin (50 μg/ml; Life Technologies, Grand Island,N.Y.). Cells were incubated at 37° C. in 5% CO₂. Cells were seeded at5.5×10⁴ cells/cm², allowed to attach overnight, and the followingmorning cells were treated with each of the test compounds. MCF-7 cellsare estrogen receptor positive and MDA-MB231 cells are estrogen receptornegative.

Cell Viability Assay:

We used an assay based on the reduction of3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide dye (MTT),which was determined according to the manufacture's protocol (Promega,Madison, Wis., USA).

Cell Proliferation Assay:

To determine cell proliferation, we measured the incorporation of5-bromo-2′-deoxyuridine (BrdU) into newly synthesized cellular DNA,following the manufacture's instructions (BD Biosciences, San Jose,Calif.).

Annexin V and Propidium Iodide (PI) Staining:

Cells were seeded at a density of 1×10⁵ cells/well and treated for 24 hwith various concentrations of each compound or equivalent volumes ofDMSO. Briefly, after incubation with the test compounds, cells weretrypsinized and stained with Annexin V-FITC (Invitrogen) and PI (0.5μg/ml). Following incubation at room temperature for 15 min in the dark,annexin V-FITC and PI fluorescence intensities were analyzed byFACScaliber (BD Bioscience). Annexin V (+)/PI (−) cells are in earlyapoptosis, annexin V (+)/PI (+) cells are in late apoptosis (secondarynecrosis), and annexin V (−)/PI (+) cells are necrotic cells.

Determination of Cell Cycle Phase Distribution (PI Incorporation Assay):

Cells were seeded in culture plates and treated for 24 h with variousconcentrations of each compound or equivalent volumes of DMSO. Aftertreatment, cells were trypsinized and fixed in 70% ethanol for 1 h onice, stained with PI (50 μg/ml) and RNase A (4 U/ml) for 30 min andsubjected to flow cytometric analysis for the determination of theirdistribution in the cell cycle phases.

The effect of the six compounds and their parent NSAIDs on six humancancer cell lines is summarized in Table 1, which shows 24-h IC₅₀s (μM)of the NSAID-derivatives in human cancer cell lines. All six compoundsshowed enhanced potency in inhibiting cell growth compared to theircorresponding conventional NSAIDs. The potency enhancement rangedbetween >6 and >63-fold (we were unable to obtain a precise IC₅₀ forconventional aspirin, given its limited solubility).

TABLE 1 24-h IC₅₀, μM BREAST COLON PANCREATIC Compound MCF-7 MDA-MB231HT29 SW480 BxPC-3 MIA-PaCa-2 Sulindac 1128 530 1173 900 489 1036Phospho-sulindac I 62 17 65 98 62 88 Ratio sulindac/PS I 18 31 18 9 8 12Phospho-sulindac II 38 18 70 73 32 92 Ratio sullindac/PS II 30 30 17 1215 11 Flurobiprofen 1433 823 1670 1216 825 1272 Phospho-Flurobiprofen 6517 80 104 34 135 Ratio flurobiprofen/PF 22 48 21 12 24 9 Ibuprofen 1229748 1554 1057 1064 1280 Phospho-Ibuprofen 79 28 82 75 53 104 Ratioibuprofen/PI 16 27 19 14 20 12 Aspirin >2000 >20003996 >2000 >2000 >2000 Glyero-phospho-aspirin I 32 199 54 169 63 303Ratio aspirin/GPA I >63 >10 74 >12 >32 >7 Glycero-phospho-aspirin II 248360 40 170 38 242 Ratio aspirin/GPA II >8 >6 100 >12 >12 >8 Note: GPA isan abbreviation for glyero-phospho-aspirin

To understand the mechanism by which these compounds inhibited cellgrowth, we evaluated in MDA-MB 231 human breast cancer cells theireffects on cell kinetics, namely cell proliferation (i.e., cellrenewal), cell death and cell cycle.

As summarized in Table 2 below, in MDA-MB231 cells, all six compounds,each used at its IC₅₀ concentration, a) inhibited cell proliferationbetween 6% and 50% compared to controls; b) induced both early and lateapoptosis, as well as necrosis; and c) inhibited the G₁ to S cell cyclephase transition. In this Table, the values for proliferation are thepercentage of the corresponding control values. Those for apoptosis andnecrosis refer to the percentage of cells in each category with respectto the entire cell population, and are to be compared to the control(vehicle only-treated) cells (uppermost row).

TABLE 2 Cell kinetic effect of six compounds Early Late Proliferationapoptosis apoptosis Necrosis Compound (% control) (%) (%) (%) ControlN/A 1.4 2.0 0.6 Phospho- 52 1.7 4.7 1.1 sulindac I Phospho- 52 5.0 23.22.6 sulindac II Phospho- 42 5.3 20.6 1.7 flurbiprofen Phospho- 50 7.443.2 5.6 ibuprofen Glycero- 94 4.8 3.2 0.3 phospho- aspirin I Glycero-52 2.4 2.2 0.4 phospho- aspirin II

Following identical methodologies as those described above for MDA-MB231cells, we determined the effect of phospho-sulindac I on the cellkinetics of SW480 and HT-29 human colon cancer cells by treating themfor 24 h with phospho-sulindac I used at its 24-h IC₅₀ concentration.Compared to untreated controls, phospho-sulindac I at a concentrationequal to its IC50 for growth at 24 h: inhibited cell proliferation by72% (from 43% in controls to 12% in phospho-sulindac I treated cells);induced apoptosis (early and late subtypes combined) by 201% (from 6.6%to 20%); induced necrosis by 1350% (from 0.2% to 2.9%) and blocked thetransition form the 01 to the S cell cycle phase

Example 5 Phospho-Sulindac I Inhibits Colon Cancer Growth In Vivo

The effect of phospho-sulindac I on tumor growth in vivo was evaluatedin two animal models of colon cancer, APC^(Min/+) mice and colon cancerxenografts in nude mice.

APC^(Min/+) Mice Study:

Min mice have a truncating mutation in the Apc gene that predisposesthem to the development of gastrointestinal tumors in the smallintestine and colon (Lipkin M, Yang K, Edelmann W, et al. Preclinicalmouse models for cancer chemoprevention studies. Ann N Y Acad Sci 1999;889:14-9.). In many important ways, this model system represents auseful (and extensively utilized) experimental system that recapitulatesthe relevant steps of colon carcinogenesis.

Eleven week-old male C57BL/6J APC^(Min/+) mice divided into four groupsof 10 mice/group were treated for 4 weeks via gavage administration asfollows: group 1 was treated with vehicle (corn oil); and group 2 wastreated with phospho-sulindac I 50 mg/kg/day. At the end of treatment,compared with vehicle-treated controls, phospho-sulindac I decreased thenumber of tumors in the small intestine by 57.2% (p<0.002) (number ofintestinal tumors in vehicle-treated group 33.6±8.7, and inphospho-sulindac I treated mice 19.4±12.0), whereas, specifically, inthe colon the reduction by phospho-sulindac I was of 61.8% (p<0.02)compared to vehicle treated mice (number of colon tumors invehicle-treated group 1.6±0.8, and in phospho-sulindac I treated mice0.6±0.5). Of note, as we have shown, conventional sulindac stimulatesthe formation of tumors in the colon of Min mice (Yang K, Fan K,Kurihara N, et al. Regional response leading to tumorigenesis aftersulindac in small and large intestine of mice with Apc mutations.Carcinogenesis 2003; 24(3):605-11). Our results document that P-S exertsa profound inhibitory effect on intestinal carcinogenesis in Min micewithout any overt signs of toxicity.

Nude Mice Xenograft Study:

Female nude mice CByJ.Cg-Foxn1 (5-6 weeks-old) were inoculatedsubcutaneously in their lower right flank with 1.5×10⁶ SW480 coloncancer cells in a volume of 100 μl (containing 50% matrigel in PBS).Seven days later, animals were randomized into two groups (8mice/group): group 1 received vehicle (1% (w/v) carboxymethylcellulose); group 2 received 50 mg/kg/day phospho-sulindac I. Alldrugs were administered in a solution of 1% (w/v) carboxymethylcellulose by gavage once daily (phospho-sulindac I and sulindacconcentrations are equimolar). Tumors were measured twice a week with adigital microcaliper, and tumor volume (TV) was calculated using theformula: TV=[L×W×(L+W/2)×0.56], where L=length and W=width of tumor.After 14 days of treatment, animals were sacrificed, and tumors wereremoved and weighed. The mean tumor weight for vehicle andphospho-sulindacl were 0.246±0.041 and 0.097±0.018 (mean±SEM),respectively, indicating a reduction in tumor weigh of 60% (p<0.05) byphospho-sulindac I. Of note, phospho-sulindac I was well tolerated,since no weight loss or other signs of toxicity were observed throughoutthe treatment period.

Synergy Between Difluoromethylornithine (DFMO) and Phospho-Sulindac

We examined the potential synergy between (DFMO) and phospho-sulindac I.A significant development in combination chemoprevention is the use ofsulindac plus DFMO to prevent colon cancer (Gerner et al. Acomprehensive strategy to combat colon cancer targeting the adenomatouspolyposis coli tumor suppressor gene. Ann N Y Acad Sci 2005;1059:97-105; Gerner E W, Meyskens F L, Jr. Polyamines and cancer: oldmolecules, new understanding. Nat Rev Cancer 2004; 4(10):781-92; GernerE W, Meyskens F L, Jr., Goldschmid S, Lance P, Pelot D. Rationale for,and design of, a clinical trial targeting polyamine metabolism for coloncancer chemoprevention. Amino Acids 2007; 33(2):189-95.). The rationalefor this combination is simple yet quite powerful: DFMO inhibits theenzyme ornithine decarboxylase, which catalyzes the rate-limiting stepin polyamine synthesis, while sulindac stimulates polyamine acetylationand export from the cell by acting on the enzyme spermidine/spermineN¹-acetyltransferase (SSAT). The end result is reduced polyamine levelsleading to suppressed growth of cancer cells. A recently published largeclinical trial demonstrated that DFMO plus sulindac reduced therecurrence of all adenomas by 69% and of advanced adenomas by 92%(Meyskens F L, McLaren C E, Pelot D, et al. Difluoromethylornithine PlusSulindac for the Prevention of Sporadic Colorectal Adenomas: ARandomized Placebo-Controlled, Double-Blind Trial. Cancer PreventionResearch Published Online First on Apr. 14, 2008 as10.1158/1940-6207.CAPR-08-0042.).

To evaluate the potential synergy between phospho-sulindac I and DFMO weused HT-29 and SW480 human colon cancer cells and employed themethodologies described above. As shown in FIG. 2A, in both HT-29 andSW480 cells DFMO 5 mM and phospho-sulindac I 40 μM each alone inhibitedmodestly cell growth at 48 h, but their combination was more effectivethan the sum of the two: a) in HT-29 cells the reductions in cell numberwere: DFMO 14%, P-S 41%, both 84%; and b) in SW480 cells: DFMO 8%, P-S45%, both 75%. Examined by isobologram (Tallarida R J, Porreca F, CowanA. Statistical analysis of drug-drug and site-site interactions withisobolograms. Life Sci 1989; 45(11):947-61), the combined effects ofDFMO and P-S on cell growth represent pharmacological synergy.

We then examined the effect of the synergy between the two compounds oncell kinetic parameters. Cell cycle analysis showed that the combinationof DFMO and phospho-sulindac I enhances the magnitude of the effect (Sphase: 13.6% for either alone is reduced to 2.9% for both). In addition,the combination of DFMO and phospho-sulindac I presented a 01 populationarrest of 88% compared to the 75% of either DFMO or phospho-sulindac Ialone (FIG. 2B). Furthermore, we examined whether DFMO increasedphospho-sulindac I-induced apoptosis in colon cancer cells. After 48 hof incubation with DFMO and phospho-sulindac I, the percentage ofapoptotic cells was 38.4%, compared to 8.7% and 16.5% for DFMO andphospho-sulindac I alone, respectively (FIG. 2C). Of note, theconcentrations of both compounds were below their IC₅₀s for cell growth.

Further evidence of the synergy between the two compounds was providedby their effect on polyamine levels. Conventional sulindac is known toreduce polyamine levels in colon cancer cells (Yerushalmi et al. Role ofpolyamines in arginine-dependent colon carcinogenesis in Apc(Min) (/+)mice. Mol Carcinog 2006; 45(10):764-73.; Choi et al. Combination of5-fluorouracil and N1,N11-diethylnorspermine markedly activatesspermidine/spermine N1-acetyltransferase expression, depletespolyamines, and synergistically induces apoptosis in colon carcinomacells. J Biol Chem 2005; 280(5):3295-304; and Basuroy and Gerner.Emerging concepts in targeting the polyamine metabolic pathway inepithelial cancer chemoprevention and chemotherapy. J Biochem (Tokyo)2006; 139(1):27-33). Phospho-sulindac I markedly diminished the levelsof spermidine (34% of control values) and spermine (9% of controlvalues) in SW480 cells, without significantly affecting those ofputrescine (91% of control values), the first polyamine in theirbiosynthetic pathway (ornithine→putrescine→spermidine→spermine).

The effect of phospho-sulindac I on polyamines is mediated, at least inpart, by activation of SSAT by phospho-sulindac I. It is known thatconventional sulindac induces SSAT activity (Babbar et al.,Cyclooxygenase-independent induction of apoptosis by sulindac sulfone ismediated by polyamines in colon cancer. J Biol Chem 2003;278(48):47762-75.). Incubation of HT-29 and SW480 cells with 85 μMphospho-sulindac I for 24 h leads to a 3- and 4.4-fold increase of SSATactivity (P<0.05 versus control), respectively. On the other hand,incubation with sulindac led to a 1.5 and 2.5-fold increase of SSATactivity (P<0.05 versus control) in HT-29 and SW480 cells, respectively.Examination of the time dependence of the induction of SSAT in SW480cells exposed to 85 μM phospho-sulindac I revealed after 12 h ofincubation, a 4-fold induction of SSAT compared to vehicle-treatedcontrols, which continued to a maximum of 5.5-fold increase after 18 hof incubation (see FIG. 4).

The Anti-Inflammatory Effect of Phospho-Sulindac I; Inhibition of NF-κBActivation

The anti-inflammatory effect of phospho-sulindac I was examined byevaluating its effect on the activation of nuclear factor-kappa B(NF-κB), a protein complex that is a transcription factor. NF-κB isfound in almost all animal cell types and is involved in cellularresponses to stimuli such as stress, cytokines, free radicals,ultraviolet irradiation, oxidized LDL, and bacterial or viral antigens(Gilmore TD (1999). “The Rel/NF-kappaB signal transduction pathway:introduction”. Oncogene 18 (49): 6842-4). NF-κB plays a key role inregulating the immune response to infection. Consistent with this role,incorrect regulation of NF-κB has been linked to cancer, inflammatoryand autoimmune diseases, septic shock, viral infection, and improperimmune development. NF-κB has also been implicated in processes ofsynaptic plasticity and memory (Albensi B C, Mattson M P (2000).“Evidence for the involvement of TNF and NF-kappaB in hippocampalsynaptic plasticity”. Synapse 35 (2): 151-9). In general, NF-κBrepresents a major molecular control of inflammation. Of additionalinterest is the modulating effect of NF-κB on cell growth andinflammation, especially in the context of cancer (Zhang Z, Rigas B.NF-kappaB, inflammation and pancreatic carcinogenesis: NF-kappaB as achemoprevention target (review). Int J Oncol 2006; 29(1):185-92; KarinM, Greten FR. NF-kappaB: linking inflammation and immunity to cancerdevelopment and progression. Nat Rev Immunol 2005; 5(10):749-59).

We investigated whether phospho-sulindac I affects the activation ofNF-κB in HT-29 human colon cancer cells, using electrophoretic mobilityshift assays. As also demonstrated in FIG. 3, we found that treatment ofHT-29 cells with phospho-sulindac I suppressed the constitutively activeNF-κB in a concentration-dependent manner. Furthermore, NF-κB activationwas rapidly induced by exposure to tumor necrosis factor alpha (TNFα) inHT-29 cells; however, 4 h pre-incubation with 80 μM phospho-sulindac Iabrogated this effect).

The Safety of Phospho-Sulindac I

We assessed the safety of phospho-sulindac I by examining itsgenotoxicity by the Ames test and its gastrointestinal and othertoxicity by performing a toxicological study in mice.

Phospho-sulindac I lacked genotoxicity, as demonstrated by amutagenicity assay performed by the BioReliance Laboratory (RockvilleMd.) that conducts such assays under high-quality standardized GoodLaboratory Pactices conditions. The mutagenic potential ofphospho-sulindac I was evaluated by measuring its ability to inducereverse mutations at selected loci of two strains of Salmonellatyphimurium in the presence and absence of S9 activation. All thesestudies were negative for genotoxicity.

Phospho-sulindac I lacked gastrointestinal toxicity in mice;gastrointestinal toxicity is the major side effect of sulindac as wellas of the entire class of NSAIDs. We evaluated in mice the potentialtoxicity of phospho-sulindac I. Three groups, each consisting of 8female C57BL/6J+/+ mice 6 wks of age, were treated for 5 days by oralgavage with equimolar amounts of phospho-sulindac I (317 mg/kg/d) orconventional sulindac (200 mg/kg/day) or vehicle. Mice were weighed attime 0 and on days 3 and 5. Mice surviving to the end of the study wereeuthanized and necropsied.

Phospho-sulindac I- and vehicle-treated mice a) maintained their weight(phospho-sulindac I=16.3±1.2→15.7±1.2; vehicle=16.1±1.0 g→15.7±1.2,mean±SD); b) showed no evidence of gastrointestinal or other toxicity;c) all were alive at the conclusion of the study and appeared healthy;and d) inspection of the heart, lungs, spleen, kidneys and liver showedno abnormalities. In contrast, sulindac-treated mice a) lost 20% oftheir weight (16.3±1.2 g→13.0±0.5 g; mean±SD); b) showed significantmortality: 75% vs. 0% for phospho-sulindac I and vehicle (5 of the 8mice died: 1 on day 2; 2 on day 3; 2 on day 4; and 1 on day 5), and c)necropsies revealed upper gastrointestinal toxicity with macroscopicallyevident gastric ulcers in 3, gastric bleeding in 1, and perforationin 1. The stomachs of sulindac-treated animals were larger than those ofthe other two groups and in some the liver appeared hyperemic.

Example 6 Phosphovalproic Acid Inhibits the Growth of Various HumanCancer Cell Lines More Potently than Conventional Valproic Acid

Valproic acid (VPA) is a in clinical use primarily as an anticonvulsantand mood-stabilizing drug, is now being extensively studied as a potentanticancer agent, especially since it was found to inhibit histonedeacetylation (Abend N S, Dlugos D J. Treatment of refractory statusepilepticus: literature review and a proposed protocol. Pediatr Neurol.2008 June; 38(6):377-90; Oki Y, Issa J P. Review: recent clinical trialsin epigenetic therapy. Rev Recent Clin Trials. 2006 May; 1(2):169-82;Barzman D H, Findling R L. Pharmacological treatment of pathologicaggression in children. Int Rev Psychiatry. 2008 April; 20(2):151-7).VPA has shown potent antitumor effects in several in vitro and in vivosystems, and encouraging results have been reported from early clinicaltrials (Duenas-Gonzalez A, Candelaria M, Perez-Plascencia C,Perez-Cardenas E, de la Cruz-Hernandez E, Herrera L A. Valproic acid asepigenetic cancer drug: preclinical, clinical and transcriptionaleffects on solid tumors. Cancer Treat Rev. 2008 May; 34(3):206-22.).

We synthesized phosphovalproic acid (phosph-VPA), a derivative of VPA,following the methodology described for phospho-sulindac I above anddetermined its effect on cell growth by determining its 24-h IC₅₀ alsoaccording to the methods described above. Conventional valproic acid wasalso studied for comparison purposes. The structure of phosphovalproicacid is:

The results summarized in Table 3, demonstrate that phosphovalproic acida) is very potent in inhibiting the growth of several human cancer celllines, and b) shows enhanced potency in inhibiting cell growth comparedto conventional VPA, with the potency enhancement ranging between 35 and245-fold.

TABLE 3 Phospho-valproic acid inhibits the growth of human cancer cells(IC₅₀, μM) Cell line VPA Phospho-VPA Fold Enhancement Breast MCF-7 1,77551 35 MDA-MB231 4,049 30 136 Colon HT-29 3,210 13 245 SW480 3,639 59 62Pancreas BxPC-3 1,680 36 47 MIA PaCa2 3,082 89 35 These values arerepresentative of two experiments, each performed in quintuplicate;results were within 10%.

The foregoing description and examples have been set forth merely toillustrate the invention and are not intended to be limiting. Sincemodifications of the disclosed embodiments incorporating the spirit andsubstance of the invention may occur to persons skilled in the art, theinvention should be construed broadly to include all variations fallingwithin the scope of the appended claims and equivalents thereof.Furthermore, the teachings and disclosures of all references citedherein are expressly incorporated in their entireties by reference.

1. A pharmaceutical composition comprising a pharmaceutically acceptableexcipient and a compound of Formula (I):

or a salt thereof, wherein A is an optionally substituted aliphatic,alicyclic, aryl, aralkyl, heteroaliphatic, heterocyclic, orheteroaromatic group; X¹ is selected from —O—, —NH—, and —S—, whereineither X¹ is —NH— or A is a radical of a non-steroidal anti-inflammatorycompound; B is an aliphatic, alicyclic, heteroaliphatic, heterocyclic,aralkyl, or heteroaromatic group optionally substituted with one or moreX² moieties, with the proviso that B is not described by

each X² is, independently, selected from hydrogen, halogen, hydroxyl,alkoxyl, —CN; an optionally substituted aliphatic, alicyclic,heteroaliphatic, heterocyclic, aryl, aralkyl, or heteroaromatic moiety;—OR^(R), —S(═O)—R^(d), —NR^(b)R^(c), —C(═O)R^(a) and —C(═O)OR^(a); n is0-2; R^(a), for each occurrence, is independently selected from hydrogenand an optionally substituted aliphatic, alicyclic, heteroaliphatic,heterocyclic, aryl, aralkyl, or heteroaromatic moiety; each of R^(b) andR^(c), for each occurrence, is independently selected from hydrogen;hydroxyl, SO₂R^(d), and an aliphatic, alicyclic, heteroaliphatic,heterocyclic, aryl, aralkyl, heteroaromatic or acyl moiety; R^(d), foreach occurrence, is independently selected from hydrogen, —N(Re)₂,aliphatic, aryl and heteroaryl, R^(e), for each occurrence, isindependently hydrogen or aliphatic; and R^(R) is an optionallysubstituted aliphatic, alicyclic, heteroaliphatic, heterocyclic, aryl,aralkyl, heteroaromatic or acyl moiety; and D is —O—P(O)(OR^(f))₂ or—O—P(OR^(g))₂, wherein each R^(f) and R^(g) is a substituted orunsubstituted alkyl of 1-6 carbon atoms.